Andresa Aparecida Berretta

Brazilian Green Propolis: Anti-Inflammatory Property by an Immunomodulatory Activity

The immunomodulatory and anti-inflammatory activities of green propolis extracts from Apis mellifera were investigated using acute and chronic inflammation models. Swiss mice were anesthetized and a cotton pellet granuloma was implanted in subcutaneous tissue. Then the mice were divided into six groups and received apyrogenic water or different propolis extracts by oral route (5 mg/kg). According to the treatment the groups were designated as E1A, E1B, E10, E11, and E12. The control group received apyrogenic water. The treatment was performed by six days when the mice were killed. The blood and the bronchoalveolar lavage (BAL) were collected to measure the leukocyte recruitment. In acute pulmonary inflammation, Balb/c mice received lipopolysaccharide (LPS) of Escherichia coli by intranasal route for three days. Concomitantly the mice received by oral route apyrogenic water (control) or E10 and E11 propolis extracts. BAL was performed to assess the inflammatory infiltrate and cytokine quantification. The results showed that the E11 extract has anti-inflammatory property in both models by the inhibition of proinflammatory cytokines and increase of anti-inflammatory cytokines suggesting an immunomodulatory activity.

Propolis Standardized Extract (EPP-AF®), an Innovative Chemically and Biologically Reproducible Pharmaceutical Compound for Treating Wounds

The aim of this study was to develop a formulation, containing the propolis standardized extract (EPP-AF®), which can assist in the healing of skin lesions. To achieve this objective the antimicrobial activity and chemical composition of the propolis extract was determined. The final product was subjected to in vitro and in vivo pre-clinical evaluation. The broth macrodilution method was used to determine the antimicrobial activity of the extracts and formulations against the microorganisms most commonly found in burns, Pseudomonas aeruginosa, Klebsiella pneumoniae, Escherichia coli, Staphylococcus aureus and Staphylococcus epidermidis. Wistar rats with puncture wounded skin were used to evaluate the wound healing properties of propolis. The results of chemical and biological characterization demonstrated the batch-to-batch reproducibility of the standardized extract which is an unprecedented result. The antimicrobial and wound healing activity of the pharmaceutical studied showed the best results when samples contain 3.6% propolis, suggesting that this is the most promising composition.

Antimicrobial Brazilian Propolis (EPP-AF) Containing Biocellulose Membranes as Promising Biomaterial for Skin Wound Healing

Among remarkable discoveries concerning propolis, such as antifungal, antiviral, and antioxidant activities, its anti-inflammatory, and mainly its antibacterial, properties deserve special attention when skin wound healing is concerned. Based on this and knowing the distinctive performance of bacterial (BC) membranes on wound healing, in this work it is proposed to demonstrate the potent antimicrobial activity and wound healing properties of a novel propolis containing biocellulose membrane. The obtained propolis/BC membrane was able to adsorb propolis not only on the surface, but also in its interstices demonstrated by scanning electron microscopy, X-ray diffraction, Fourier transform infrared (FT-IR) spectroscopy, and thermogravidimetric assays. Additionally, the polyphenolic compounds determination and the prominent antibacterial activity in the membrane are demonstrated to be dose dependent, supporting the possibility of obtaining propolis/BC membranes at the desired concentrations, taking into consideration its application and its skin residence time. Finally, it could be suggested that propolis/BC membrane may favor tissue repair in less time and more effectively in contaminated wounds.

Molecular Characterization of Propolis-Induced Cell Death in Saccharomyces cerevisiae

ABSTRACT Propolis, a natural product of plant resins, is used by the bees to seal holes in their honeycombs and protect the hive entrance. However, propolis has also been used in folk medicine for centuries. Here, we apply the power of Saccharomyces cerevisiae as a model organism for studies of genetics, cell biology, and genomics to determine how propolis affects fungi at the cellular level. Propolis is able to induce an apoptosis cell death response. However, increased exposure to propolis provides a corresponding increase in the necrosis response. We showed that cytochrome c but not endonuclease G (Nuc1p) is involved in propolis-mediated cell death in S. cerevisiae. We also observed that the metacaspase YCA1 gene is important for propolis-mediated cell death. To elucidate the gene functions that may be required for propolis sensitivity in eukaryotes, the full collection of about 4,800 haploid S. cerevisiae deletion strains was screened for propolis sensitivity. We were able to identify 138 deletion strains that have different degrees of propolis sensitivity compared to the corresponding wild-type strains. Systems biology revealed enrichment for genes involved in the mitochondrial electron transport chain, vacuolar acidification, negative regulation of transcription from RNA polymerase II promoter, regulation of macroautophagy associated with protein targeting to vacuoles, and cellular response to starvation. Validation studies indicated that propolis sensitivity is dependent on the mitochondrial function and that vacuolar acidification and autophagy are important for yeast cell death caused by propolis.

Identification of the cell targets important for propolis-induced cell death in Candida albicans

Candida albicans is the most common fungal pathogen of humans, forming both commensal and opportunistic pathogenic interactions, causing a variety of skin and soft tissue infections in healthy people. In immunocompromised patients C. albicans can result in invasive, systemic infections that are associated with a high incidence of mortality. Propolis is a complex mixture of several resinous substances which are collected from plants by bees. Here, we demonstrated the fungicidal activity of propolis against all three morphogenetic types of C. albicans and that propolis-induced cell death was mediated via metacaspase and Ras signaling. To identify genes that were involved in propolis tolerance, we screened ~800 C. albicans homozygous deletion mutants for decreased tolerance to propolis. Fifty-one mutant strains were identified as being hypersensitive to propolis including seventeen genes involved in cell adhesion, biofilm formation, filamentous growth, phenotypic switching and pathogenesis (HST7, GIN4, VPS34, HOG1, ISW2, SUV3, MDS3, HDA2, KAR3, YHB1, NUP85, CDC10, MNN9, ACE2, FKH2, and SNF5). We validated these results by showing that propolis inhibited the transition from yeast-like to hyphal growth. Propolis was shown to contain compounds that conferred fluorescent properties to C. albicans cells. Moreover, we have shown that a topical pharmaceutical preparation, based upon propolis, was able to control C. albicans infections in a mouse model for vulvovaginal candidiasis. Our results strongly indicate that propolis could be used as a strategy for controlling candidiasis.

Assessment of the Mutagenic Activity of Extracts of Brazilian Propolis in Topical Pharmaceutical Formulations on Mammalian Cells In Vitro and In Vivo

Propolis possesses various biological activities such as antibacterial, antifungal, anti-inflammatory, anesthetic and antioxidant properties. A topically applied product based on Brazilian green propolis was developed for the treatment of burns. For such substance to be used more safely in future clinical applications, the present study evaluated the mutagenic potential of topical formulations supplemented with green propolis extract (1.2, 2.4 and 3.6%) based on the analysis of chromosomal aberrations and of micronuclei. In the in vitro studies, 3-h pulse (G1 phase of the cell cycle) and continuous (20 h) treatments were performed. In the in vivo assessment, the animals were injured on the back and then submitted to acute (24 h), subacute (7 days) and subchronic (30 days) treatments consisting of daily dermal applications of gels containing different concentrations of propolis. Similar frequencies of chromosomal aberrations were observed for cultures submitted to 3-h pulse and continuous treatment with gels containing different propolis concentrations and cultures not submitted to any treatment. However, in the continuous treatment cultures treated with the 3.6% propolis gel presented significantly lower mitotic indices than the negative control. No statistically significant differences in the frequencies of micronuclei were observed between animals treated with gels containing different concentrations of propolis and the negative control for the three treatment times. Under the present conditions, topical formulations containing different concentrations of green propolis used for the treatment of burns showed no mutagenic effect in either test system, but 3.6% propolis gel was found to be cytotoxic in the in vitro test.

Draft Genome Sequence of Komagataeibacter rhaeticus Strain AF1, a High Producer of Cellulose, Isolated from Kombucha Tea.

ABSTRACT Here, we present the draft genome sequence of Komagatabaeicter rhaeticus strain AF1, which was isolated from Kombucha tea and is capable of producing high levels of cellulose.

The Inhibition of Inflammasome by Brazilian Propolis (EPP-AF)

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-1β and 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 effective in reducing the IL-1β secretion in mouse macrophages and this reduction was correlated with a decrease in activation of the protease caspase-1. Furthermore, we found that the extract at a concentration of 30 μg/mL was not toxic to the cells even after a 18-hour treatment. Altogether, these data indicate that Brazilian green propolis (EPP-AF) extract has a role in regulating the inflammasomes.

Effect of the Copaifera langsdorffii Desf. Leaf Extract on the Ethylene Glycol-Induced Nephrolithiasis in Rats

The potential of the Copaifera langsdorffii leaves extract to prevent stone formation was analyzed by means of an ethylene glycol (EG) animal model of nephrolithiasis and an in vitro crystallization assay. Different doses of the C. langsdorffii leaves extract were administered to rats treated with EG. Urine biochemical parameters were quantified. CaOx deposits count and analysis of osteopontin expression were conducted on kidneys fixed in formalin. The in vitro assay was performed by turbidimetry. Phytochemical analyses of the extract were accomplished by HPLC-UV-DAD, and several compounds were isolated. C. langsdorffii leaf extract was able to avoid stone formation. The number of deposits was 50.30 ± 31.29 at the higher extract dose, compared to the value of 179.5 ± 45.96 achieved with the EG control. Significantly lower oxalate levels and OPN expression and increased citrate levels were observed after extract administration. In the in vitro assay, the extract diluted the formed crystals. Phytochemical analyses showed that the extract is rich in phenolic compounds that are capable of preventing stone formation. Thus, on the basis of our results, we suggest that the C. langsdorffii leaf extract has potential application in the prevention of kidney stone formation.

Evaluation of Mucoadhesive Gels with Propolis (EPP-AF) in Preclinical Treatment of Candidiasis Vulvovaginal Infection

Vulvovaginal candidiasis is the second cause of vaginal infection in the USA. Clinical treatment of C. albicans infections is routinely performed with polyenes and azole derivatives. However, these drugs are responsible for undesirable side effects and toxicity. In addition, C. albicans azole and echinocandin resistance has been described. Propolis is a bee product traditionally used due to its antimicrobial, anti-inflammatory, and other properties. Therefore, the present work aimed to evaluate different propolis presentations in order to evaluate their in vitro and in vivo efficacy. The methodologies involved antifungal evaluation, chemical analysis, and the effects of the rheological and mucoadhesive properties of propolis based gels. The obtained results demonstrated the fungicide action of propolis extracts against all three morphotypes (yeast, pseudohyphae, and hyphae) studied. The highest level of fungal cytotoxicity was reached at 6–8 hours of propolis cell incubation. Among the based gel formulations developed, the rheological and mucoadhesive results suggest that propolis based carbopol (CP1%) and chitosan gels were the most pseudoplastic ones. CP1% was the most mucoadhesive preparation, and all of them presented low thixotropy. Results of in vivo efficacy demonstrated that propolis based gels present antifungal action similar to clotrimazole cream, suggesting that future clinical studies should be performed.