Juliana Hori

Prosthetic stomatitis is often associated with microbial colonization of the acrylic base of the prostheses, favored by their poor hygiene and the high level of <em>Candida</em> spp. in the oral cavity. <em>Candida albicans</em> is the fungal species most commonly associated with this oral pathology, usually treated with antifungals. The indiscriminate use of these drugs can promote resistance of these species and side effects to individuals. Thus, it is desirable to promote health through natural or alternative therapies, such as the use of herbal compounds. Studies have shown promising results on the action of some herbal compounds in the treatment of infections, such as pomegranate, sisal and propolis. The objective of this study will be to evaluate <em>in vivo</em>, the safety and tolerability of phytotherapy gel based on propolis extracts and associations with pomegranate and sisal and its effectiveness in reducing the prevalence of <em&gt...

Copyright © 2013 Juliana I. Hori et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Propolis extracts have gained the attention of consumers and researchers due to their unique chemical compositions and functional properties such as its anti-inflammatory activity. Recently, it was described a complex that is also important in inflammatory processes, named inflammasome. The inflammasomes are a large molecular platform formed in the cell cytosol in response to stress signals, toxins, and microbial infections. Once activated, the inflammasome induces caspase-1, which in turn induces the processing of inflammatory cytokines such as IL-1

Copyright © 2015 Felipe Galeti Miguel et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Chamomile is a medicinal plant, which presents several biological effects, especially the anti-inflammatory effect. One of the compounds related to this effect is apigenin, a flavonoid that is mostly found in its glycosylated form, apigenin-7-glucoside (APG), in natural sources. However, the affectivity and safety of this glycoside have not been well explored for topical application. In this context, the aim of this work was to develop and validate a reversed-phase high-performance liquid chromatography (RP-HPLC-DAD) method to quantify APG in chamomile preparations. Additionally, the safety and the anti-inflammatory potential

We used immunocytochemical and fluorescence assays to investigate the subcellular location of the protein encoded by Down syndrome critical region gene 2 (DSCR2) in transfected cells. It was previously suggested that DSCR2 is located in the plasma membrane as an integral membrane protein. Interestingly, we observed this protein in the endoplasmic reticulum (ER) of cells. We also studied whether the truncated forms of DSCR2 showed different subcellular distributions. Our observations indicate that DSCR2 probably is not inserted into the membrane of the endoplasmic reticulum since the fragments lacking the predicted transmembrane (TM) helices remained associated with the ER. Our analyses suggest that, although DSCR2 is associated with the endoplasmic reticulum, it is not an integral membrane protein and it is maintained on the cytoplasmic side of the ER by indirect interaction with the ER membrane or with another protein.

Propolis is a natural product produced by bees that is primarily used in complementary and alternative medicine and has anti-inflammatory, antibacterial, antiviral, and antitumoral biological properties. Some studies have reported the beneficial effects of propolis in models of allergic asthma. In a previous study, our group showed that green propolis treatment reduced airway inflammation and mucus secretion in an ovalbumin (OVA)-induced asthma model and resulted in increased regulatory T cells (Treg) and polymorphonuclear myeloid-derived suppressor cells (PMN-MDSC) frequencies in the lungs, two leukocyte populations that have immunosuppressive functions. In this study, we evaluated the anti-inflammatory effects of artepillin C (ArtC), the major compound of green propolis, in the context of allergic airway inflammation. Our results show that ArtC induces in vitro differentiation of Treg cells and monocytic MDSC (M-MDSC). Furthermore, in an OVA-induced asthma model, ArtC treatment re...

PURPOSE We therefore evaluated the analgesic, anti-inflammatory and toxicological effects of indomethacin incorporated into mesoporous silica nanoparticles (IND+NP). METHODS Nociception was evaluated by the formalin assay. The anti-inflammatory potential was assessed by cell migration and paw edema assays, modulation of nitric oxide and cytokines (IL-6, IL-10 and TNF-α) by macrophages production. Toxicity was evaluated in peritoneal macrophages and by the locomotion assay and assessment of gastric injuries, presence of occult blood and hepatic and renal markers. RESULTS IND+NP reduced nociception during phases 1 by 53% and 2 by 79% of the formalin assay and the influx of peritoneal cells by 94%, indicating an analgesic and anti-inflammatory effect more efficiently than indomethacin alone. Indomethacin, but not IND+NP, caused macroscopic gastric injuries, the presence of fecal occult blood, and an increase of ALT levels. In the paw edema assay, IND+NP reduced edema by 21%. IND+NP has no effect on the LPS-induced production of nitric oxide, IL-6, IL-10 and TNF-α on no cytotoxic concentrations. CONCLUSIONS The incorporation of indomethacin into mesoporous silica nanoparticles effectively increased the activity of the drug observed in the formalin and cell migration assays and prevented the gastric and hepatic damage associated with its use.

ETHNOPHARMACOLOGICAL RELEVANCE Propolis is a natural product produced by honeybees used as a medicine at least to 300 BC. In the last decades, several studies showed biological and pharmacological properties of propolis, witch scientifically explains the empirical use for centuries. The anti-inflammatory activity of propolis with the purpose to reduce Th2 inflammation has been evaluated in allergic asthma. However, it remains to be determined how propolis negatively regulates the immune response after allergen re-exposure. AIM OF THE STUDY We hypothesized that the anti-inflammatory activity of propolis is dependent on the induction of myeloid derived suppressor cells (MDSC) and regulatory T cells. MATERIALS AND METHODS To assess this hypothesis, we used an ovalbumin-induced asthma model to evaluate the effect of EPP-AF® dry extract from Brazilian green propolis. RESULTS Propolis treatment decreased pulmonary inflammation and mucus production as well as eosinophils and IL-5 in the broncoalveolar lavage. Propolis enhanced also in vitro differentiation and in vivo frequency of lung MDSC and CD4+Foxp3+ regulatory T cells. CONCLUSIONS Together these results confirm the immunomodulatory potential of propolis during sensitization and challenge with allergen. In addition, the collecting findings show, for the first time, that propolis increases the frequency of MDSC and CD4+Foxp3+ regulatory T cells in the lungs, and suggest that it could be use as target for development of new immunotherapy or adjuvant immunotherapy for asthma.

Antimalarials have demonstrated beneficial effects in Systemic Lupus Erithematosus and Rheumatoid Arthritis. However, the mechanisms and the molecular players targeted by these drugs remain obscure. Although hydroxychloroquine (HCQ) is a known ion channel inhibitor, this property has not been linked to its anti-inflammatory effects. We aimed to study whether HCQ inhibits pro-inflammatory ion channels. Electrophysiology experiments demonstrated that HCQ inhibited Ca(++)-activated K(+) conductance in THP-1 macrophages in a dose-dependent manner. In macrophages, ATP-induced K(+) efflux plays a key role in activating the NLRP3 inflammasome. ATP-induced IL-1beta secretion was controlled by the KCa1.1 inhibitor iberiotoxin. NS1619 and NS309 (KCa1.1 and KCa3.1 activators respectively) induced the secretion of IL-1beta. This effect was inhibited by HCQ and also by iberiotoxin and clotrimazol (KCa3.1 inhibitor), arguing against off-target effect. In vitro, HCQ inhibited IL-1beta and caspase ...

The Cell Wall Integrity (CWI) pathway is the primary signaling cascade that controls the de novo synthesis of the fungal cell wall and in Saccharomyces cerevisiae, this event is highly dependent on the RLM1 transcription factor. Here, we investigated the function of RlmA in the fungal pathogen Aspergillus fumigatus We show that the ΔrlmA strain exhibits an altered cell wall organization in addition to defects related to vegetative growth and tolerance to cell wall-perturbing agents. A genetic analysis indicated that rlmA is positioned downstream of the pkcA and mpkA genes in the CWI pathway. As a consequence, rlmA loss-of-function leads to the altered expression of genes encoding cell wall-related proteins. RlmA positively regulates the phosphorylation of MpkA and is induced at both protein and transcriptional levels during cell wall stress. The rlmA was also involved in tolerance to oxidative damage and transcriptional regulation of genes related to oxidative stress adaptation. Mor...

Tityus serrulatus sting causes thousands of deaths annually worldwide. T. serrulatus-envenomed victims exhibit local or systemic reaction that culminates in pulmonary oedema, potentially leading to death. However, the molecular mechanisms underlying T. serrulatus venom (TsV) activity remain unknown. Here we show that TsV triggers NLRP3 inflammasome activation via K(+) efflux. Mechanistically, TsV triggers lung-resident cells to release PGE2, which induces IL-1β production via E prostanoid receptor 2/4-cAMP-PKA-NFκB-dependent mechanisms. IL-1β/IL-1R actions account for oedema and neutrophil recruitment to the lungs, leading to TsV-induced mortality. Inflammasome activation triggers LTB4 production and further PGE2 via IL-1β/IL-1R signalling. Activation of LTB4-BLT1/2 pathway decreases cAMP generation, controlling TsV-induced inflammation. Exogenous administration confirms LTB4 anti-inflammatory activity and abrogates TsV-induced mortality. These results suggest that the balance betwe...

Chamomile is a medicinal plant, which presents several biological effects, especially the anti-inflammatory effect. One of the compounds related to this effect is apigenin, a flavonoid that is mostly found in its glycosylated form, apigenin-7-glucoside (APG), in natural sources. However, the affectivity and safety of this glycoside have not been well explored for topical application. In this context, the aim of this work was to develop and validate a reversed-phase high-performance liquid chromatography (RP-HPLC-DAD) method to quantify APG in chamomile preparations. Additionally, the safety and the anti-inflammatory potential of this flavonoid were verified. The RP-HPLC-DAD method was developed and validated with linearity at 24.0–36.0 μg/mL range (r=0.9994). Intra- and interday precision (RSD) were 0.27–2.66% and accuracy was 98.27–101.21%. The validated method was applied in the analysis of chamomile flower heads, glycolic extract, and Kamillen cream, supporting the method applica...

Aspergillus fumigatus is an opportunistic pathogenic fungus able to infect immunocompromised patients, eventually causing disseminated infections that are difficult to control and lead to high mortality rates. It is important to understand how the signalling pathways that regulate these factors involved in virulence are orchestrated. Protein phosphatases are central to numerous signal transduction pathways. Here we characterize A. fumigatus protein phosphatase 2A SitA, the S. cerevisiae Sit4p homologue. The sitA gene is not an essential gene and we were able to construct an A. fumigatus null mutant. The ΔsitA strain had decreased MpkA phosphorylation levels, was more sensitive to cell wall damaging agents, had increased β-1,3-glucan and chitin, was impaired in biofilm formation, and had decreased protein kinase C activity. The ΔsitA strain is more sensitive to several metals and ions such as MnCl2, CaCl2, and LiCl, however, it is more resistant to ZnSO4. The ΔsitA strain was avirule...

Legionella pneumophila, the etiological agent of Legionnaires' disease, triggers activation of multiple innate immune pathways that lead to the restriction of bacterial replication in vivo. Despite the critical role for MyD88 in infection clearance, the receptors and mechanisms responsible for MyD88-mediated pulmonary bacterial clearance are still unclear. Here, we used flagellin mutants of L. pneumophila, which bypass the NAIP5/NLRC4-mediated restriction of bacterial replication, to assess the receptors involved in MyD88-mediated pulmonary bacterial clearance. By systematically comparing pulmonary clearance of L. pneumophila in C57BL/6 MyD88(-/-), TLR2(-/-), TLR3(-/-), TLR4(-/-), TLR9(-/-), IL-1R(-/-), and IL-18(-/-) mice, we found that, while the knockout of a single Toll-like receptor or interleukin 18 resulted only in minor impairment of bacterial clearance, deficiency in the interleukin 1 (IL-1) receptor led to a significant impairment. IL-1/MyD88-mediated pulmonary bacteri...

Legionella pneumophila is an intracellular bacterium that was evolutionarily selected to survive in freshwater environments by infecting free-living unicellular protozoa. Once humans inhale contaminated water droplets, the bacteria reach the pulmonary alveoli where they are phagocytized by resident alveolar macrophages. Depending on host immunity and bacterial virulence genes, the infection may progress to an acute pneumonia called Legionnaires' disease, which can be fatal. Of note, an effective immune response is critical to the outcome of the human infection. These clinical observations highlight the importance of animal models of pulmonary infection for in vivo investigation of bacterial pathogenesis and host responses. In this chapter we provide detailed protocols for intranasal infection of mouse with L. pneumophila.

Tuberculosis is a major threat to human health. The high disease burden remains unaffected and the appearance of extremely drug-resistant strains in different parts of the world argues in favor of the urgent need for a new effective vaccine. One of the promising candidates is heat-shock protein 65 when used as a genetic vaccine (DNAhsp65). Nonetheless, there are substantial data indicating that BCG, the only available anti-TB vaccine for clinical use, provides other important beneficial effects in immunized infants. We compared the protective efficacy of BCG and Hsp65 antigens in mice using different strategies: i) BCG, single dose subcutaneously; ii) naked DNAhsp65, four doses, intramuscularly; iii) liposomes containing DNAhsp65, single dose, intranasally; iv) microspheres containing DNAhsp65 or rHsp65, single dose, intramuscularly; and v) prime-boost with subcutaneous BCG and intramuscular DNAhsp65. All the immunization protocols were able to protect mice against infection, with special benefits provided by DNAhsp65 in liposomes and prime-boost strategies. Among the immunization protocols tested, liposomes containing DNAhsp65 represent the most promising strategy for the development of a new anti-TB vaccine.

Propolis extracts have gained the attention of consumers and researchers due to their unique chemical compositions and functional properties such as its anti-inflammatory activity. Recently, it was described a complex that is also important in inflammatory processes, named inflammasome. The inflammasomes are a large molecular platform formed in the cell cytosol in response to stress signals, toxins, and microbial infections. Once activated, the inflammasome induces caspase-1, which in turn induces the processing of inflammatory cytokines such as IL-1βand IL-18. So, to understand inflammasomes regulation becomes crucial to treat several disorders including autoinflammatory diseases. Since green propolis extracts are able to regulate inflammatory pathways, this work purpose was to investigate if this extract could also act on inflammasomes regulation. First, the extract was characterized and it demonstrated the presence of important compounds, especially Artepillin C. This extract was...

Propolis extracts have gained the attention of consumers and researchers due to their unique chemical compositions and functional properties such as its anti-inammatory activity. Recently, it was described a complex that is also important in inammatory processes, named inammasome. e inammasomes are a large molecular platform formed in the cell cytosol in response to stress signals, toxins, and microbial infections. Once activated, the inammasome induces caspase-, which in turn induces the processing of inammatory cytokines such as IL-and IL-. So, to understand inammasomes regulation becomes crucial to treat several disorders including autoinammatory diseases. Since green propolis extracts are able to regulate inammatory pathways, this work purpose was to investigate if this extract could also act on inammasomes regulation. First, the extract was characterized and it demonstrated the presence of important compounds, especially Artepillin C. is extract was eective in reducing the IL-secretion in mouse macrophages and this reduction was correlated with a decrease in activation of the protease caspase-. Furthermore, we found that the extract at a concentration of g/mL was not toxic to the cells even aer a-hour treatment. Altogether, these data indicate that Brazilian green propolis (EPP-AF) extract has a role in regulating the inammasomes.

The control and treatment of Leishmaniasis, a neglected and infectious disease affecting approximately 12 million people worldwide, are challenging. Leishmania parasites multiply intracellularly within macrophages located in deep skin and in visceral tissues, and the currently employed treatments for this disease are subject to significant drawbacks, such as resistance and toxicity. Thus, the search for new Leishmaniasis treatments is compulsory, and Ocotea duckei Vattimo, a plant-derived product from the biodiverse Brazilian flora, may be a promising new treatment for this disease. In this regard, the aim of this work was to develop and characterize a delivery system based on solid lipid nanoparticles (SLN) that contain the liposoluble lignan fraction (LF) of Ocotea duckei Vattimo, which targets the Leishma-nia phagolysosome of infected macrophages. LF-loaded SLNs were obtained via the hot microemulsion method, and their physical and chemical properties were comprehensively assessed using PCS, AFM, SEM, FT-IR, DSC, HPLC, kinetic drug release studies, and biological assays. The size of the developed delivery system was 218.85 ± 14.2 nm, its zeta potential was −30 mV and its entrapment efficiency (EE%) was high (the EEs% of YAN [yangambin] and EPI-YAN [epi-yangambin] markers were 94.21 ± 0.40% and 94.20 ± 0.00%, respectively). Microscopy, FT-IR and DSC assays confirmed that the delivery system was nanosized and indicated a core–shell encapsulation model, which corroborated the measured kinetics of drug release. The total in vitro release rates of YAN and EPI-YAN in buffer (with sink conditions attained) were 29.6 ± 8.3% and 34.3 ± 8.9%, respectively, via diffusion through the cellulose acetate membrane of the SLN over a period of 4 h. After 24 h, the release rates of both markers reached approximately 45%, suggesting a sustained pattern of release. Mathematical modeling indicated that both markers, YAN and EPI-YAN, followed matrix diffusion-based release kinetics (Higuchi's model) with an estimated diffusion coefficient (D) of 1.3.10 −6 cm 2 /s. The LF-loaded SLNs were non-toxic to murine macrophages (20–80 g mL −1 range) and exerted a prominent anti-leishmanial effect (20 g mL −1). These data suggest this new and well-characterized lipid nanoparticle delivery system safely and effectively kills Leishmania and warrants further clinical investigation.

The Cell Wall Integrity (CWI) pathway is the primary signaling cascade that controls the de novo synthesis of the fungal cell wall, and in Saccharomyces cerevisiae this event is highly dependent on the RLM1 transcription factor. Here, we investigated the function of RlmA in the fungal pathogen Aspergillus fumigatus. We show that the DrlmA strain exhibits an altered cell wall organization in addition to defects related to vegetative growth and tolerance to cell wall-perturbing agents. A genetic analysis indicated that rlmA is positioned downstream of the pkcA and mpkA genes in the CWI pathway. As a consequence, rlmA loss-of-function leads to the altered expression of genes encoding cell wall-related proteins. RlmA positively regulates the phosphorylation of MpkA and is induced at both protein and transcriptional levels during cell wall stress. The rlmA was also involved in tolerance to oxidative damage and transcriptional regulation of genes related to oxidative stress adaptation. Moreover, the DrlmA strain had attenuated virulence in a neutropenic murine model of invasive pulmonary aspergillosis. Our results suggest that RlmA functions as a transcription factor in the A. fumigatus CWI pathway, acting downstream of PkcA-MpkA signaling and contributing to the virulence of this fungus. KEYWORDS