[Show abstract][Hide abstract] ABSTRACT: Propolis is a complex resinous mixture collected by bees, with high medicinal, historical and economic value. The nutraceutical and pharmacological benefits of propolis have been extensively explored in several fields of medicine as an important resource for prevention and treatment of oral and systemic diseases. A relatively new type of propolis, named red propolis (in Brazil, Brazilian Red Propolis - BRP), has been arousing attention for the promising pharmacological properties of some of its isolated compounds (vestitol, neovestitol, quercetin, medicarpin, formononetin, etc). Due to a distinct chemical composition, BRP and its isolated compounds (mainly isoflavones) affect a wide range of biological targets and could have an impact against numerous diseases as an antimicrobial, anti-inflammatory and immunomodulatory, antioxidant and antiproliferative agent. In this review, we comprehensively address the main aspects related to BRP bioprospection, chemistry and therapeutic potential. Further information is provided on mechanisms of action discovered thus far as well as clinical use in humans and regulatory aspects. As of now, BRP and its isolated molecules remain a fascinating topic for further research and application in biomedical areas and dentistry.
Full-text · Article · Mar 2016 · European Journal of Medicinal Chemistry
[Show abstract][Hide abstract] ABSTRACT: Although previous studies suggested an anti-inflammatory property of Brazilian red propolis (BRP), the mechanisms involved in the anti-inflammatory effects of BRP and its activity on macrophages were still not elucidated. This study aimed to evaluate whether BRP attenuates the inflammatory effect of LPS on macrophages and to investigate its underlying mechanisms. BRP was added to RAW 264.7 murine macrophages after activation with LPS. NO production, cell viability, cytokines profile were evaluated. Activation of inflammatory signaling pathways and macrophage polarization were determined by RT-qPCR and Western blot. BRP at 50 μg/ml inhibited NO production by 78% without affecting cell viability. Cd80 and Cd86 were upregulated whereas mrc1 was down regulated by BRP indicating macrophage polarization at M1. BRP attenuated the production of pro-inflammatory mediators IL-12, GM-CSF, IFN-Ɣ, IL-1β in cell supernatants although levels of TNF- α and IL-6 were slightly increased after BRP treatment. Levels of IL-4, IL-10 and TGF-β were also reduced by BRP. BRP significantly reduced the up-regulation promoted by LPS of transcription of genes in inflammatory signaling (Pdk1, Pak1, Nfkb1, Mtcp1, Gsk3b, Fos and Elk1) and of Il1β and Il1f9 (fold-change rate > 5), which were further confirmed by the inhibition of NF-κB and MAPK signaling pathways. Furthermore, the upstream adaptor MyD88 adaptor-like (Mal), also known as TIRAP, involved in TLR2 and TLR4 signaling, was down- regulated in BRP treated LPS-activated macrophages. Given that BRP inhibited multiple signaling pathways in macrophages involved in the inflammatory process activated by LPS, our data indicated that BRP is a noteworthy food-source for the discovery of new bioactive compounds and a potential candidate to attenuate exhacerbated inflammatory diseases.
[Show abstract][Hide abstract] ABSTRACT: Fractionation of geopropolis from Melipona scutellaris, guided by antiproliferative activity against two colon cancer cell lines (COLO205 and KM12), led to the isolation of two new cinnamic acid esters, mammea-type coumarins 5,7-dihydroxy-6-(3-methyl-2-butenyl)-8-(4-cinnamoyl-3-methyl-1-oxobutyl)-4-propyl-coumarin (1) and 5,7-dihydroxy-6-(4-cinnamoyl-3-methyl-1-oxobutyl)-4-phenylcoumarin (2), along with five known coumarins, mammeigin (3), hydroxymammeigin (4), mammeisin (5), cinnamoyloxy-mammeisin (6), and mammein (7), and the prenylated benzophenone ent-nemorosone (8). Among the isolated compounds, 5 and 7 showed the highest cell growth inhibition against COLO205 (GI50 9.7 and 10.7 µM, respectively) and KM12 (GI50 12.0 and 10.9 µM, respectively). The presence of these compounds suggests that plants of Clusiaceae family, especially the genera Kielmeyera and Clusia, are likely to be major sources of geopropolis produced by M. scutellaris.
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to investigate the in vitro anti-inflammatory activity of Malva sylvestris extract (MSE) and fractions in a co-culture model of cells infected by Aggregatibacter actinomycetemcomitans. In addition, we evaluated the phytochemical content in the extract and fractions of M. sylvestris and demonstrated that polyphenols were the most frequent group in all samples studied. An in vitro dual-chamber model to mimic the periodontal structure was developed using a monolayer of epithelial keratinocytes (OBA-9) and a subepithelial layer of fibroblasts (HGF-1). The invasive periodontopathogen A. actinomycetemcomitans (D7S-1) was applied to migrate through the cell layers and induce the synthesis of immune factors and cytokines in the host cells. In an attempt to analyze the antimicrobial properties of MSE and fractions, a susceptibility test was carried out. The extract (MIC 175 μg/mL, MBC 500μg/mL) and chloroform fraction (MIC 150 μg/mL, MBC 250 μg/mL) were found to have inhibitory activity. The extract and all fractions were assessed using a cytotoxicity test and results showed that concentrations under 100 μg/mL did not significantly reduce cell viability compared to the control group (p > 0.05, viability > 90%). In order to analyze the inflammatory response, transcriptional factors and cytokines were quantified in the supernatant released from the cells. The chloroform fraction was the most effective in reducing the bacterial colonization (p< 0.05) and controlling inflammatory mediators, and promoted the down-regulation of genes including IL-1beta, IL-6, IL-10, CD14, PTGS, MMP-1 and FOS as well as the reduction of the IL-1beta, IL-6, IL-8 and GM-CSF protein levels (p< 0.05). Malva sylvestris and its chloroform fraction minimized the A. actinomycetemcomitans infection and inflammation processes in oral human cells by a putative pathway that involves important cytokines and receptors. Therefore, this natural product may be considered as a successful dual anti-inflammatory-antimicrobial candidate.
[Show abstract][Hide abstract] ABSTRACT: Background:
Subantimicrobial dose doxycycline (SDD) has been used as an adjunct in periodontal treatment due to its matrix metalloproteinase inhibition properties. Although the benefits of SDD therapy, such as improvement in the parameters of periodontal probing depth and clinical attachment level, have been proven in multiple clinical studies, the comprehension of other biological mechanisms of action on periodontitis remains poorly investigated. Therefore, this animal model study evaluated the effects of SDD monotherapy on the expressions of the following key pro-inflammatory genes: Proteinase-Actived Receptor-2 (PAR2), Tumor Necrosis Factor alpha (TNF-α), Interleukin-17 (IL-17), and. IL-1β.
Male Wistar rats were randomly assigned to: A) Control group: no ligature-induced periodontitis and no treatment; B) Ligature group: ligature-induced periodontitis and placebo treatment and C) Ligature + doxycycline group: ligature-induced periodontitis and SDD treatment. After the experimental time, animals were sacrificed and Reverse Transcriptase-Polymerase Chain Reaction was performed to analyze the mRNA expression of IL-1β, IL-17, TNF-α, and PAR2 in gingival tissue samples. Histological analyses were performed on the furcation region and mesial gingiva of mandibular first molars to measure periodontal bone loss and collagen content.
SDD administration significantly downregulated PAR2, IL-17, TNF-α, and IL-1β mRNA expressions (p<0.05). In addition, SDD treatment was accompanied by lower rates of alveolar bone loss (p<0.05) and by maintenance of the amount of gingival collagen fibers.
These findings reveal new perspectives regarding SDD efficacy since it can be partially related to pro-inflammatory gene expression modulation, even considering PAR2 and IL-17, which has not been investigated thus far.
No preview · Article · Oct 2015 · Journal of Periodontology
[Show abstract][Hide abstract] ABSTRACT: Abstract Nearly 20 million tons of winery by-products, with many biological activities, are discarded each year in the world. The extraction of bioactive compounds from Chenin Blanc, Petit Verdot, and Syrah grape by-products, produced in the semi-arid region in Brazil, was optimized by a Central Composite Rotatable Design. The phenolic compounds profile, antioxidant capacity against synthetic free radicals (DPPH and ABTS), reactive oxygen species (ROS; peroxyl radical, superoxide radical, hypochlorous acid), cytotoxicity assay (MTT) and quantification of TNF-α production in RAW 264.7 cells were conducted. Gallic acid, syringic acid, procyanidins B1 and B2, catechin, epicatechin, epicatechin gallate, quercetin 3-β-d-glucoside, delfinidin 3-glucoside, peonidin 3-O-glucoside, and malvidin 3-glucoside were the main phenolic compounds identified. In general, rachis showed higher antioxidant capacity than pomace extract, especially for Chenin Blanc. All extracts showed low cytotoxicity against RAW 264.7 cells and Petit Verdot pomace suppressed TNF-α liberation in vitro. Therefore, these winery by-products can be considered good sources of bioactive compounds, with great potential for application in the food and pharmaceutical industries.
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to evaluate the gastroprotective activity of ethanolic extract of geopropolis (EEGP) from Melipona scutellaris and to investigate the possible mechanisms of action. The gastroprotective activity of the EEGP was evaluated using model ulcer induced by ethanol. To elucidate the possible mechanisms of action, we investigated the involvement of the nonprotein sulfhydryl (NP-SH) groups, nitric oxide and prostaglandins. In addition, the antisecretory activity of EEGP was also evaluated by pylorus ligated model. The EEGP orally administrated (300 mg/kg) reduced the ulcerative lesions induced by the ethanol (). Regarding the mechanism of action, the prior administration of nitric oxide and prostaglandins antagonists suppressed the activity of gastroprotective EEGP (). On the other hand the gastroprotective activity of EEGP was kept in the group pretreated with the antagonist of the NP-SH groups; furthermore the antisecretory activity was not significant (). These results support the alternative medicine use of geopropolis as gastroprotective and the activities observed show to be related to nitric oxide and prostaglandins production.
Preview · Article · May 2015 · Evidence-based Complementary and Alternative Medicine
[Show abstract][Hide abstract] ABSTRACT: A collection of tagged deletion mutant strains was created in Streptococcus mutans UA159 to facilitate investigation of the aciduric capability of this oral pathogen. Gene-specific barcoded deletions were attempted in 1432 open reading frames (representing 73% of the genome), and resulted in the isolation of 1112 strains (56% coverage) carrying deletions in distinct non-essential genes. Since S. mutans virulence is predicated upon the ability of the organism to survive an acidic pH environment, form biofilms on tooth surfaces, and out-compete other oral microflora, we assayed individual mutant strains for the relative fitness of the deletion strain, compared to the parent strain, under acidic and oxidative stress conditions, as well as for their ability to form biofilms in glucose- or sucrose-containing medium. Our studies revealed a total of 51 deletion strains with defects in both aciduricity and biofilm formation. We have also identified 49 strains whose gene deletion confers sensitivity to oxidative damage and deficiencies in biofilm formation. We demonstrate the ability to examine competitive fitness of mutant organisms using the barcode tags incorporated into each deletion strain to examine the representation of a particular strain in a population. Co-cultures of deletion strains were grown either in vitro in a chemostat to steady-state values of pH 7 and 5 or in vivo in an animal model for oral infection. Taken together, this data represents a mechanism for assessing the virulence capacity of this pathogenic microorganism and a resource for identifying future targets for drug intervention to promote healthy oral microflora. This article is protected by copyright. All rights reserved.
This article is protected by copyright. All rights reserved.
[Show abstract][Hide abstract] ABSTRACT: In oral biofilms, two of the major environmental challenges encountered by the dental pathogen Streptococcus mutans are acid and oxidative stresses. Previously, we showed that the S. mutans transcriptional regulators SpxA1 and SpxA2 (formerly SpxA and SpxB, respectively) are involved in stress survival by activating the expression of classic oxidative stress genes such as dpr, nox, sodA and tpx. We reasoned that some of the uncharacterized genes under SpxA1/A2 control are potentially involved in oxidative stress management. Therefore, the goal of this study was to use Spx-regulated genes as a tool to identify novel oxidative stress genes in S. mutans. Quantitative real-time PCR was used to evaluate the responses of ten Spx-regulated genes during H2O2 stress in the parent and Δspx strains. Transcription activation of the H2O2-induced genes (8 out of 10) was strongly dependent on SpxA1 and, to a lesser extent, SpxA2. In vitro transcription assays revealed that one or both Spx proteins directly regulate three of these genes. The gene encoding the FeoB ferrous permease was slightly repressed by H2O2 but constitutively induced in strains lacking SpxA1. Nine genes were selected for downstream mutational analysis but inactivation of smu127, encoding a subunit of the acetoin dehydrogenase was apparently lethal. In vitro and in vivo characterization of the viable mutants indicated that, in addition to the transcriptional activation of reducing and antioxidant pathways, Spx performs an important role in iron homeostasis by regulating the intracellular availability of free iron. In particular, inactivation of the genes encoding the Fe-S biogenesis SUF system and the previously characterized iron-binding protein Dpr resulted in impaired growth under different oxidative stress conditions, increased sensitivity to iron and lower infectivity in rats. These results serve as an entryway into the characterization of novel genes and pathways that allow S. mutans to cope with oxidative stress.
[Show abstract][Hide abstract] ABSTRACT: Dental caries remains the most prevalent and costly oral infectious disease worldwide. Several methods have been employed to prevent this biofilm-dependent disease, including the use of essential oils (EOs). In this systematic review, we discuss the antibacterial activity of EOs and their isolated constituents in view of a potential applicability in novel dental formulations. Seven databases were systematically searched for clinical trials, in situ, in vivo and in vitro studies addressing the topic published up to date. Most of the knowledge in the literature is based on in vitro studies assessing the effects of EOs on caries-related streptococci (mainly Streptococcus mutans) and lactobacilli, and on a limited number of clinical trials. The most promising species with antibacterial potential against cariogenic bacteria are: Achillea ligustica, Baccharis dracunculifolia, Croton cajucara, Cryptomeria japonica, Coriandrum sativum, Eugenia caryophyllata, Lippia sidoides, Ocimum americanum, and Rosmarinus officinalis. In some cases, the major phytochemical compounds determine the biological properties of EOs. Menthol and eugenol were considered outstanding compounds demonstrating an antibacterial potential. Only L. sidoides mouthwash (1%) has shown clinical antimicrobial effects against oral pathogens thus far. This review suggests avenues for further non-clinical and clinical studies with the most promising EOs and their isolated constituents bioprospected worldwide.
[Show abstract][Hide abstract] ABSTRACT: The essential oils (EO) and bioactive fractions (BF) from Aloysia gratissima, Baccharis dracunculifolia, Coriandrum sativum, Cyperus articulatus, and Lippia sidoides were proven to have strong antimicrobial activity on planktonic microorganisms; however, little is known about their effects on the morphology or viability of oral biofilms. Previously, we determined the EO/fractions with the best antimicrobial activity against Streptococcus mutans and Candida spp. In this report, we used a confocal analysis to investigate the effect of these EO and BF on the morphology of S. mutans biofilms (thickness, biovolume, and architecture) and on the metabolic viability of C. albicans biofilms. The analysis of intact treated S. mutans biofilms showed no statistical difference for thickness in all groups compared to the control. However, a significant reduction in the biovolume of extracellular polysaccharides and bacteria was observed for A. gratissima and L. sidoides groups, indicating that these BF disrupt biofilm integrity and may have created porosity in the biofilm. This phenomenon could potentially result in a weakened structure and affect biofilm dynamics. Finally, C. sativum EO drastically affected C. albicans viability when compared to the control. These results highlight the promising antimicrobial activity of these plant species and support future translational research on the treatment of dental caries and oral candidiasis.
Full-text · Article · Mar 2015 · Evidence-based Complementary and Alternative Medicine
[Show abstract][Hide abstract] ABSTRACT: Objective: recent data have shown that Brazilian red propolis presents immunemodulatory properties, which led us to elucidate the mechanisms involved in the anti-inflammatory properties of one of its components, Neovestitol
Method: neovestitol was obtained from ethanolic extract of Brazilian red propolis after bio-guided fractionation coupled with GC-MS analysis. Different concentrations of Neovestitol were added to RAW 264.7 macrophages after activation with LPS. After 48 hours, NO production and cell viability were determined. Production of TNF-α, IL1β, TGF-β, IL-4, IL-6, IL-10, IL-12, GM-CSF, IFN-Ɣ and expression of genes related to cytokines production, nitric oxide , PI3K-AKT and signal transduction pathways were evaluated by ELISA and RT-qPCR, respectively. Differences were determined by one-way ANOVA followed by Tukey-Kramer
Result: Neovestitol inhibited NO production by 63% without affecting cell viability at the concentration of 60 µg/ml when compared with control cells (treated with vehicle) (p<0.05). Levels of GM-CSF, IFN- Ɣ, IL-1 β, IL-4, TNF- α and IL-6 in cell supernatant were reduced and of IL-10 increased with the addition of neovestitol (p<0.05). Neovestitol induced alteration in the expression of 43 genes. Genes involved in Toll-like response; nitric oxide sintetase production (all types), Nf-KB and Il-1 signaling pathways such as Bad, Chuk, Elk1, Ifnb1, Il1b, Lif, Calm1, Capns1, Egr1, Nox1, Scd2 and Scd3 were negatively regulated by neovestitol (fold-change rate > 5; p<0.05). Furthermore, its role in inhibition of leukocyte transmigration was suggested by inhibition of expression of Nox1 and in the regulation of fatty acid associated genes such as Scd2 e Scd3
Conclusion: Neovestitol may represent a new antiinflammatory agent by inhibiting signaling pathways and consequently reducing the production of pro-inflammatory factors and by interfering in leukocyte transmigration
[Show abstract][Hide abstract] ABSTRACT: Objective: to elucidate the mechanisms involved in the anti-inflammatory properties of Brazilian red propolis
Method: different concentrations of ethanolic extract of Brazilian red propolis (EEBRP) were added to RAW 264.7 macrophages after activation with LPS, and NO production and cell viability determined. Production of TNF-α, IL1β, TGF-β, IL-4, IL-6, IL-10, IL-12, GM-CSF, IFN-Ɣ and expression of genes related to cytokines production and nitric oxide , PI3K-AKT and signal transduction pathway were evaluated by ELISA and RT-qPCR, respectively. Differences were determined by one-way ANOVA followed by Tukey-Kramer
Result: EEBRP inhibited NO production by 69% without affecting cell viability at the concentration of 50 µg/ml when compared with control cells (treated with vehicle) (p<0.05). Levels of IL-12, GM-CSF, IFN-Ɣ, IL-1β, IL-4, IL-10 and TGF-β in cell supernatant were reduced and of TNF- α and IL-6 increased with the addition of EEBRP (p<0.05). EEBRP induced alteration in expression of 57 genes. Genes involved in Toll-like response; oxide nitric sintetase production (all types), Nf-KB and MAPK signaling pathways such as Tirap, Pdk1, Pak1, Nfkb1, Mtcp1, Gsk3b, Fos, Elk1, Il1β, Il1f9 were negatively regulated by EEBRP(fold-change rate > 5; p<0.05)
Conclusion: EEBRP presents antiinflammatory properties by altering signaling pathways leading to reduction in production of pro-inflammatory factors.
[Show abstract][Hide abstract] ABSTRACT: The aim of the present study was to perform an in vitro analysis of the antimicrobial and antiproliferative potential of an extract from Anadenanthera colubrina (Vell.) Brenan (angico) and chemically characterize the crude extract. Antimicrobial action was evaluated based on the minimum inhibitory concentration (MIC), minimum bactericidal/fungicidal concentration, and the inhibition of formation to oral biofilm. Cell morphology was determined through scanning electron microscopy (SEM). Six strains of tumor cells were used for the determination of antiproliferative potential. The extract demonstrated strong antifungal activity against Candida albicans ATCC 18804 (MIC = 0.031 mg/mL), with similar activity found regarding the ethyl acetate fraction. The extract and active fraction also demonstrated the capacity to inhibit the formation of Candida albicans to oral biofilm after 48 hours, with median values equal to or greater than the control group, but the difference did not achieve statistical significance (P > 0.05). SEM revealed alterations in the cell morphology of the yeast. Regarding antiproliferative activity, the extract demonstrated cytostatic potential in all strains tested. The present findings suggest strong antifungal potential for Anadenanthera colubrina (Vell.) Brenan as well as a tendency toward diminishing the growth of human tumor cells.
Full-text · Article · Jun 2014 · Evidence-based Complementary and Alternative Medicine