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Camila Faria de Amorim Pereira,1,* Michelle Nonato de Oliveira Melo,1,* Vania Emerich Bucco de Campos,2 Ivania Paiva Pereira,1 Adriana Passos Oliveira,1 Mariana Souza Rocha,1 João Vitor da Costa Batista,3,4 Valter Paes de Almeida,5 Irailson Thierry Monchak,5 Eduardo Ricci-Júnior,6 Rafael Garrett,7 Aline Gabrielle Alves Carvalho,7 Jane Manfron,5 Stephan Baumgartner,3,8,9 Carla Holandino1,3 1Multidisciplinary Laboratory of Pharmaceutical Sciences, Faculty of Pharmacy, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil; 2Department of Pharmacy, Universidade do Estado do Rio de Janeiro, Rio de Janeiro, Brazil; 3Society for Cancer Research, Hiscia Institute, Arlesheim, Switzerland; 4Department of Pharmaceutical Sciences, Division of Pharmaceutical Technology, University of Basel, Basel, Switzerland; 5Postgraduate Program in Pharmaceutical Sciences, Universidade Estadual de Ponta Grossa, Ponta Grossa, Paraná, Brazil; 6Galenic Development Laboratory (LADEG), Department of Drugs and Medicines, Faculty of Pharmacy, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil; 7Metabolomics Laboratory, Chemistry Institute, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil; 8Institute of Integrative Medicine, University of Witten/Herdecke, Herdecke, Germany; 9Institute of Complementary and Integrative Medicine, University of Bern, Bern, Switzerland*These authors contributed equally to this workCorrespondence: Carla Holandino, Multidisciplinary Laboratory of Pharmaceutical Sciences, Universidade Federal do Rio de Janeiro, Faculty of Pharmacy, Block B basement, Room 34, 373, Carlos Chagas Filho Avenue, Cidade Universitária, Rio de Janeiro, RJ, 21941-902, Brazil, Email cholandino@gmail.com Stephan Baumgartner, Institute of Complementary and Integrative Medicine, University of Bern, Bern, Switzerland, Email St.Baumgartner@vfk.chBackground and Purpose: Natural products are potential sources of anticancer components. Among various species, the lipophilic extract of the Viscum album subsp. austriacum (Wiesb.) Vollm. (VALE) has shown promising therapeutic potential. The present work aimed to qualify the plant source and characterize the extract’s chemical profile. In addition, a self-nanoemulsifying drug delivery system (SNEDDS) containing VALE (SNEDDS-VALE) was developed.Methods: V. album subsp. austriacum histochemistry was performed, and the chemical profile of VALE was analyzed by GC-MS. After the SNEEDS-VALE development, its morphology was visualized by transmission electron microscopy (TEM), while its stability was evaluated by the average droplet size, polydispersity index (PdI) and pH. Lastly, SNEDDS-VALE chemical stability was evaluated by LC-DAD-MS.Results: The histochemical analysis showed the presence of lipophilic compounds in the leaves and stems. The major compound in the VALE was oleanolic acid, followed by lupeol acetate and ursolic acid. SNEDDS was composed of medium chain triglyceride and Kolliphor® RH 40 (PEG-40 hydrogenated castor oil). A homogeneous, isotropic and stable nanoemulsion was obtained, with an average size of 36.87 ± 1.04 nm and PdI of 0.14 ± 0.02, for 14 weeks.Conclusion: This is the first histochemistry analysis of V. album subsp. austriacum growing on Pinus sylvestris L. which provided detailed information regarding its lipophilic compounds. A homogeneous, isotropic and stable SNEDDS-VALE was obtained to improve the low water solubility of VALE. Further, in vitro and in vivo experiments should be performed, in order to evaluate the antitumoral potential of SNEDDS-VALE. Keywords: Viscum album subsp. austriacum, mistletoe, lipophilic extract, oleanolic acid, SNEDDS

Anderson Nogueira Mendes,1 Lívia Alves Filgueiras,1 Monica Regina Pimentel Siqueira,2,3 Gleyce Moreno Barbosa,2 Carla Holandino,2 Davyson de Lima Moreira,3 José Carlos Pinto,4 Marcio Nele5 1Department of Biophysics and Physiology, Federal University of Piauí, Teresina, Piauí, 2School of Pharmacy, Federal University of Rio Janeiro, 3Natural Products Department, Institute of Pharmaceutical Technology, Farmanguinhos, Oswaldo Cruz Foundation, Rio de Janeiro, 4Chemical Engineering Program – COPPE, Federal University of Rio de Janeiro, Rio de Janeiro 5School of Chemistry, Federal University of Rio Janeiro, Rio de Janeiro, Brazil Abstract: This study aimed to synthesize and characterize nanoparticles (NPs) of poly(methyl methacrylate) (PMMA) and evaluate their ability to incorporate plant extracts with antitumor activity and low dissolution in aqueous media. The extract used was n-hexane partition of the methanol extract of Piper cabralanum (PCA-HEX). PMMA NPs were obtained using the miniemulsion method, which was able to encapsulate almost 100% of PCA-HEX. The synthesized polymeric particles presented with a size of 200 nm and a negative charge. Cytotoxicity tests by MTT and trypan blue assays showed that NPs without PCA-HEX did not kill leukemic cells (K562 cells). NPs containing PCA-HEX were able to enhance cell death when compared to pure extract. The results showed that PMMA NPs could be useful as a drug delivery system as they can enhance the antitumor activity of the PCA-HEX extract by more than 20-fold. PMMA NPs containing plant extracts with antitumor activities may be an alternative to control the evolution of diseases such as leukemia. Keywords: Piper, Piperaceae, nanoparticles, miniemulsion, PMMA, K562 cells, leukemia

J Oral Pathol Med (2010) 39: 753-760 Background: There are several kinds of oral soft tissue lesions that are common manifestations observed in human immunodeficiency virus (HIV)-infected children; for example, linear gingival erythema (LGE) that is a distinctive fiery red band along the margin of the gingivae. The etiology and pathogenesis of LGE are questionable, but a candidal origin has been suggested. Proteases are key virulence attributes produced by a variety of pathogenic fungi, including Candida. The objective of the present study is to identify the protease production in Candida species including, C. albicans (n = 5), C. dubliniensis (n = 1) and C. tropicalis (n = 1), isolated directly from typical LGE lesions observed in six HIV-positive children, and also to test the effect of a serine protease inhibitor on the interaction of Candida spp. and epithelial cells in vitro. Methods: The ability of Candida strains to release proteases in the culture supernatant fluids was visualized by gelatin-SDS-PAGE. Gel strips containing 30-fold concentrated supernatant (1.5 x 10(8) yeasts) were incubated at 37°C for 48 h in 50 mM sodium phosphate buffer, pH 5.5. The concentrated supernatants were also incubated with fibronectin, laminin, immunoglobulin G, bovine serum albumin and human serum albumin. The effect of serine protease inhibitor on the interaction of Candida spp. and epithelial cells (MA 104) was measured after pre-treatment of fungi with the inhibitor (phenylmethylsulphonyl fluoride, PMSF). Results: All the extracellular proteases were completely inhibited by PMSF, identifying these activities as serine-type proteases. Interestingly, a common 62-kDa serine protease was observed in all Candida strains. The culture supernatants, rich in serine protease activities, cleaved several soluble proteinaceous substrates. Additionally, we demonstrated that pre-treatment of C. albicans, C. dubliniensis and C. tropicalis with PMSF diminished the interaction with epithelial cells. Conclusions: Collectively, our results show that Candida spp. isolated from LGE lesions produced and secreted serine proteases and these enzymes may be involved in the initial colonization events. [ABSTRACT FROM AUTHOR]

Objective: Pharmacopoeias regulate the manufacture of potentised pharmaceutical preparations used in different branches of complementary and integrative medicine. The physicochemical properties and biological activity of these preparations are often investigated in preclinical research, yet no guidelines for experimental research currently exist in this area. The present PrePoP guidelines aim to provide recommendations to promote high-quality, statistically sound, and reproducible preclinical research on potentised preparations., Methods: Input was gathered from researchers nominated by the relevant scientific societies using a simplified Delphi consensus approach covering the most relevant aspects of basic research methodology in the field including appropriate controls, sample preparation and handling, and statistics. After three rounds of feedback, a consensus was finally reached on the most important aspects and considerations for conducting high-quality research on potentised preparations., Results: We present a series of recommendations on a range of topics including experimental controls, system stability, blinding and randomisation, environmental influences, and procedures for the preparation of potentised samples and controls, and we address some specific challenges of this research field., Conclusion: This expert consensus process resulted in a robust set of methodological guidelines for research on potentised preparations and provides a valuable framework that will inform and improve the quality of subsequent research in this emerging field., Please Cite This Article as: Tournier AL, Bonamin LV, Buchheim-Schmidt S, Cartwright S, Dombrowsky C, Doesburg P, Holandino C, Kokornaczyk MO, van de Kraats EB, López-Carvallo JA, Nandy P, Mazón-Suástegui JM, Mirzajani F, Poitevin B, Scherr C, Thieves K, Würtenberger S, Baumgartner S. Scientific guidelines for preclinical research on potentised preparations manufactured according to current pharmacopoeias-the PrePoP guidelines. J Integr Med. 2024; 22(5): 533-544., (Copyright © 2024 Shanghai Yueyang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine. Published by Elsevier B.V. All rights reserved.)

Background:  Coronavirus disease 2019 (COVID-19), a disease caused by the SARS-CoV-2 coronavirus, was declared a pandemic in March 2020, posing significant challenges globally. Homeopathy has historical relevance in epidemic management. In response, the government of the state of Santa Catarina, Brazil, distributed Camphora 1M as a potential prophylactic intervention for COVID-19., Objective:  This study aimed to investigate the possible effects of Camphora 1M as an adjunctive prophylactic measure in managing COVID-19, focusing on mortality and hospitalization rates, during the period April 28 to July 31, 2020, within designated COVID-19 in-patient units in Santa Catarina., Methods:  An ecological study design was applied to this epidemiological research. Five case municipalities (Itajaí, Atalanta, Entre Rios, Rio do Campo, Trombudo Central) were compared with five control municipalities (São José, Galvão, Pedras Grandes, Grão-Pará, Ascurra)., Results:  No statistically significant differences were observed in predictor variables between municipalities that received Camphora 1M and the respective controls. Similarly, no statistically significant differences were observed in outcomes: deaths ( p  = 0.879), hospitalized cases ( p  = 0.537), daily ward admissions ( p  = 0.730) and ICU admissions ( p  = 0.072)., Conclusion:  For the first wave of the pandemic in the state of Santa Catarina, Brazil, city-wide distribution of Camphora 1M was not associated with reduced numbers, severity or mortality among the population hospitalized in designated public hospitals for COVID-19., Competing Interests: None declared., (Faculty of Homeopathy. This article is published by Thieme.)

Aconitum napellus L. is a popular medicinal plant extensively used in homeopathy. This article provides detailed morphology and microscopy, including the anatomical and histochemical features of the herb, to aid authentication and quality control. In cross-section, the root in secondary growth shows the phloem surrounded by pericyclic fibers and a well-developed xylem. The stem is irregular in outline, displaying unicellular trichomes and many free collateral vascular bundles encircling the pith. The leaf is dorsiventral, hypostomatic with anomocytic and anisocytic stomata, and shows non-glandular trichomes. The floral parts are characterized by uniseriate epidermises, homogeneous mesophyll, anomocytic stomata on the abaxial surface, trichomes, and oval pollen grains. The tissue fragments in powdered herbs show these characteristics and have numerous starch grains with thimble-shaped, linear or star-shaped hilum. The detailed macroscopic and microscopic analysis provided in this study can help in the authentication and quality control of A. napellus raw materials. RESEARCH HIGHLIGHTS: Key anatomical, micromorphological, and microchemical features of Aconitum napellus are described. The results of the study can support the taxonomy of the genus Aconitum. Morphological standardization of the species reported here is helpful in the quality control of this herb., (© 2023 Wiley Periodicals LLC.)

Introduction: Larch oleoresin has been described regarding several biological activities and medicinal applications, such as wound healing and treatment of ulcers, but little is known about its chemical composition., Material and Methods: Eight oleoresins from Larix decidua Mill. obtained from four companies and one adulterated control were therefore investigated to determine their content of essential oils and to verify possible differences in their composition in relation to the harvest and manufacturing processes. Essential oils (EOs) were isolated by distillation and the yield was analysed., Results and Discussion: The yield of EO varied among all samples. The yield of the pure larch samples covered a range of 7.8% to 15.5%. A higher yield (19.0%) was observed for adulterated control, which contained oleoresins from different Pinaceae trees. Age of samples had no impact on yield. However, there was a significant statistical variation (p<0.05) in the yields of the mid-summer oleoresins (>10%) compared to early or late summer (<10%), emphasising the importance of the time of collection. Samples were subsequently analysed by GC-MS. EO samples confirmed the presence of various chemical classes, such as monoterpenes, sesquiterpenes, and diterpenes. α-pinene was the compound with the highest concentrations (>50%), followed by β-pinene (>6%), D-limonene (>2.5%), α-terpineol (>0.9%), β-myrcene (>0.2%), and 3-carene (>0.05%). Samples were grouped using multivariate data analysis (MVDA) with respect to the chemical variation between the oleoresins' EOs. The resulting four clusters were named low (low yield obtained for the samples), mixed (mixed oleoresin from different Pinaceae species, adulteration control), old (old oleoresin kept in the institute), and normal (other oleoresins) samples, each presenting distinct chemical biomarkers. There were considerable differences between site and time of collection. Essential oil yield did not always meet requirements as defined by the German Homeopathic Pharmacopoeia. In addition, adulterated or aged samples could be identified as compared to pure and fresh larch oleoresins., Conclusion: We conclude that larch oleoresin used for pharmaceutical applications has to be carefully analysed and standardised to guarantee reproducible product quality., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Batista, Melo, Holandino, Maier, Huwyler, Baumgartner and Boylan.)

The genus Viscum comprises a large number of semi-parasitic shrubs popularly known as Mistletoe. The Viscum species grow in many countries of Europe, Africa and Asia with different popular uses in ornamentation, foods and medicine. Many studies about Viscum have been done over the last years focusing on biological activities and chemical composition of the aqueous extracts, mainly related to anthroposophical medicines. However, it is known that non-aqueous preparations, as alcoholic extracts, have demonstrated different biological activities that are species-and host tree-dependent. Considering the potential of these alcoholic extracts, a scoping review was conducted using data from three online databases: PubMed, Scopus and Embase. Inclusion criteria consisted of the in vitro, in vivo, ex vivo, clinical and chemical studies of alcoholic extracts from Viscum species. The present review summarized 124 original publications about fourteen Viscum species. Viscum album , Viscum articulatum and Viscum coloratum were the main studied species. Alcoholic extracts demonstrated hypotensive, anticancer, antimicrobial, analgesic and anti-inflammatory capabilities, among other biological activities. Flavonoids, phenolic acids and terpenoids represented 48%, 24% and 11% of the total identified compounds, respectively. This review contributes to the knowledge of alcoholic preparations of the Viscum species and points out the lack of clinical studies concerning these different extracts.

Viscum album L. (Santalaceae) is an important medicinal plant traditionally used to treat several diseases, including cancer therapy. This paper provides detailed morpho-anatomical characteristics of the leaves, stems and berries of Viscum album subsp. album growing as hemi-parasite on the branches of Malus domestica (Suckow) Borkh. (Rosaceae) to aid species identification and botanical characterization. Additionally, for the first time, microchemical analyses of all tissues and Energy Dispersive X-Ray Spectroscopy analyses of the calcium oxalate crystals are provided for the first time. The plant features leathery presents green leaves with parallel veins, small yellow unisexual flowers in 3-flowered cymes, and the dioecious inflorescences usually consist of three flowers, with female flowers generating white fleshy berries, in which a seed is embedded in the mucilaginous mesocarp, normally containing two embryos. Anatomically, the analyzed leaves were isobilateral and amphistomatic, and showed straight anticlinal epidermal cell walls, thick cuticles with epicuticular wax crystalloids, and paracytic stomata. The midrib is flat on both sides and has a single vascular bundle, whereas the strongly shortened petiole is concave-convex in shape and contains five bundles. The stems show a primary structure with a ring of nine vascular bundles enclosing the pith. Calcium oxalate druses and cubic and quadrangular prisms were observed in different plant parts. The results of this study provide new microscopy information that can help in the authentication of mistletoe raw materials., (© 2023. The Author(s).)

Viscum album L., popularly known as mistletoe, is well known for its anti-cancer properties, and the pharmaceutical application of hydroalcoholic dry extracts is still limited due to its low solubility in aqueous media, and physicochemical instability. The Pluronic® F127 is an amphiphilic polymer, which permits the solubilization of lipophilic and hydrophilic compounds. In this investigation, physicochemical features of hydrogel containing V. album dry extract (VADE-loaded-hydrogel) were performed by: dynamic light scattering (DLS), thermogravimetric analysis (TGA), differential scanning calorimetry (DSC), and transmission electron microscopy (TEM). VADE-loaded-hydrogel presented nanometer-size micelles with volume distribution ranging from 10.58 nm to 246.7 nm, and a polydispersity index of 0.441. The sample thermal analyses (TG and DSC) showed similar decomposition curves; however, the thermal events indicated an increase in thermal stability in relation to the presence of the extract. In addition to these interesting pharmaceutical features, IC50 values of 333.40 µg/mL and >1000 µg/mL were obtained when tumor (SCC-25) and non-tumor (L929) cells were incubated with VADE-loaded-hydrogel, respectively. The optical and ultrastructural cellular analysis confirmed the tumor selectivity since the following alterations were detected only in SCC-25 cells: disorganization of plasmatic membrane; an increase of cytoplasmatic vacuole size; alteration in the cristae mitochondrial shape; and generation of amorphous cellular material. These results emphasize the promising antitumoral potential of VADE-loaded-hydrogel as an herbal drug delivery system via in vitro assays.

Viscum album is a semi-parasitic plant used for over one hundred years in complementary cancer therapy. The main commercial drugs used in cancer patients' treatment are derived from the aqueous V. album extracts, whose cytotoxic potential is mostly attributed to the aqueous soluble antitumoral metabolites. On the counterpart, ethanol solvents must be used to obtain V. album mother tinctures. This methodology permits better solubilization of phenolic compounds, among others, which present antitumoral bioactivity. Recently, the metabolomics approach revealed the influence of the host tree on the V. album subspecies differentiation. To increase the scientific information about the chemical differences related to the host trees and to clarify the seasonal influences, in this study, the metabolome of 50  V. album mother tinctures from three subspecies ( abietis , album , austriacum ) and five host trees ( Malus domestica , Quercus sp. , Ulmus carpinifolia , Pinus sylvestris , Abies alba ) was evaluated using summer and winter plant harvests. The in vitro cytotoxic activities were investigated in breast cancer cells (MDA-MB-231) and immortalized normal human keratinocytes (HaCaT). The summer V. album mother tinctures presented higher cytotoxic activity than winter ones. Among the summer samples, those prepared with V. album subsp. album were more cytotoxic than V. album subsp. abietis and subsp. V. album subsp. austriacum . The V. album harvested from Quercus petraea and Abies alba inhibited the key-glycolytic enzymes: hexokinase (HK), phosphofructokinase (PFK), pyruvate kinase (PK). This activity was related to a reduction in glucose uptake and lactate production, which were host-tree-time-dose-dependent. The untargeted metabolomic approach was able to discriminate the mother tinctures according to respective botanical classes and harvest season. A total of 188 metabolites were annotated under positive and negative modes. Fourteen compounds were responsible for the samples differentiation, and, to the best of our knowledge, eight were described in the Viscum album species for the first time. Our study shows the interruption of the Warburg effect as a novel antitumoral mechanism triggered by V. album mother tinctures, which is related to their metabolite profile. These results bring scientific evidence that encourages the use of V. album mother tinctures as a natural product for cancer therapy., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Melo, Ochioni, Zancan, Oliveira, Grazi, Garrett, Holandino and Baumgartner.)

Background: Recent experimental results supporting the dynamization process show modification in the characteristics of solid mixtures., Objective: The present work aims to evaluate the physicochemical properties of metallic zinc and lactose, evidencing the interactions between all chemical components presented in dynamized solid mixtures by analytical techniques., Methods: Mixtures of zinc and lactose (1:9 w/w) were successively triturated at the same proportion according to the Brazilian Homeopathic Pharmacopoeia, receiving the designation of 10 -1 - 10 -6 (1dH - 6dH). All samples were submitted to the following characterization techniques: Atomic Absorption Spectrometry (AAS), Scanning Electron Microscopy (SEM), X-ray Diffraction (XRD), Differential Scanning Calorimetry (DSC), Thermogravimetry (TG), and Raman Spectroscopy (RS)., Results: AAS results detected 97.0% of zinc in the raw material, and the triturated zinc lactose system ( ZnMet ) presented mean values similar to those expected for the physical mixtures: i.e., 9.94%, 1.23%, and 0.11% in the three first proportions (10 -1 , 10 -2 , 10 -3 ), respectively. SEM images showed particle size reduction due to the trituration process. The XRD assays of ZnMet 10 -3 and 10 -6 indicated peak changes at 12.3° and 43.26°, probably associated with modifications of inter-atomic crystalline spacing. The thermal analysis results of dynamized samples suggest modifications in the chemical interaction between zinc and lactose induced by the physical forces applied. RS experiments showed variation in vibration frequencies due to the dynamization procedure, in which marked ZnMet 10 -6 spectral modifications were detected at 357, 477, 1086 and 1142 cm -1 , and in the wavelength range 860-920 cm -1 ., Conclusion: These results highlight the importance of applying suitable characterization methods to improve our understanding of the properties of homeopathic solid mixtures, whereas the uses of sensitive tools evidence the influence of trituration on the crystalline properties and in the enthalpy variation of dynamized samples., Competing Interests: Disclosure The authors report no conflicts of interest in this work., (Faculty of Homeopathy. This article is published by Thieme.)

Malignant ulcerating wounds or neoplastic lesions are a considerable burden for patients suffering from advanced cancer. These wounds have no effective treatment and are very difficult to manage. The present review summarizes evidence in support of a hypothesis put forward in anthroposophic medicine, which suggests a beneficial role of resin from the species Larix decidua Mill. [Pinaceae] for treating such wounds. A systematic search strategy was performed using the databases PubMed, EMBASE and SciFinder. The included publications described the chemical composition of this species, as well as in vitro , in vivo , and ex vivo experiments using plant extracts and isolated compounds. The results show that among the phytochemical classes, terpenoids were the major components of this species, especially in the resin. The summarized biological experiments revealed antimicrobial, antioxidant and anti-inflammatory effects, with promising potential for the extracts and isolated compounds. However, the molecular mechanisms and toxicological effects are as of yet not conclusively evaluated. From the data of our study, we can conclude that L. decidua might indeed have a promising potential for the treatment of malignant wounds, but definitive information that can prove its effectiveness is still lacking. We therefore suggest that future efforts should be dedicated to the evaluation of L. decidua resin's therapeutic use considering its antiseptic action and proposed wound healing properties., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Batista, Uecker, Holandino, Boylan, Maier, Huwyler and Baumgartner.)

Context and Objective: Periodontitis and type 2 diabetes (T2D) are chronic diseases generally treated with conventional therapies alone. The aim of this study was to compare the effects of homeopathy as an adjunct to conventional periodontal therapy in individuals with periodontitis and T2D., Design: 85 individuals, age between 35 and 70 years, of both genders, participated in this randomized study; 70 patients were from the Institute of Endocrinology and Diabetes of Rio de Janeiro, Brazil. They were divided into 2 groups: G1, individuals with periodontitis without systemic conditions; and G-2, individuals with periodontitis and T2D. Both groups received homeopathic treatment and were evaluated in clinical and laboratory examinations. The medication used was chosen based on the similarity principle: Berberis 6CH, Mercurius Solubilis/Belladona /Hepar Sulfur and a Pyrogenium 200CH biotherapic. Medications were prescribed in diluted low ultra-diluted concentration doses for all signs and symptoms, while biotherapics were used for chronic stimulation., Setting: The study was performed in Brazil by university research professors of homeopathy and periodontics., Results: Both groups showed significant clinical and laboratory improvements during the study from baseline to 1 year with reductions in total cholesterol (total-C), triglycerides, glucose, glycated hemoglobin (A1cHb), uric acid and C-reactive protein (CRP). Statistical and descriptive analyses were performed. For most parameters, G1 performed better than G2 (P < .05)., Conclusion: Homeopathy as an adjunct to periodontal treatment improves local and systemic clinics and can provide better health conditions for patients with or without T2D.

Viscum album L. ( Santalaceae ), also known as European mistletoe, is a semi-parasitic plant that grows on different host trees. Our group recently demonstrated the antitumoral activity of ethanolic V. album extracts in vitro, depending on the dose and the host tree, V. album ssp abietis from Abies alba being the most active extract. The goal of this work focused on the development of a new topical formulation containing V. album extracts, evaluation of in vitro toxicity and ex vivo skin permeation assays. The Poloxamer 407 hydrogel containing 5% of dry (VA&#95;DEH) or aqueous (VA&#95;AEH) extract presented dermal compatible pH and microbiological stability for 180 days. The hydrogels flow curve presented a non-linear relation, characteristic of non-Newtonian fluids, and the mean viscosity for the VA&#95;DEH and VA&#95;AEH was 372.5 ± 7.78 and 331.0 ± 2.83 Pa.s, respectively, being statistically different (Welch's t test; p < 0.01). Additionally, WST-1 in vitro assays revealed a dose-dependent toxicity for both formulations and VA&#95;DEH presented a higher activity than the VA&#95;AEH. The promising cytotoxic potential of VA&#95;DEH lead to the ex vivo skin permeation assay with 2.73 ± 0.19 µg/cm 2 of chlorogenic acid, which permeated at 8 h, showing a transdermal potential. These in vitro results support the idea that VA&#95;DEH is a novel promising candidate for mistletoe therapy. Therefore, further in vivo and pre-clinical experiments should be performed to evaluate the safety and efficacy of this new dermic delivery system.

Viscum album L., commonly known as European mistletoe, is a hemi-parasitic plant of the Santalaceae family. The in vitro and in vivo effects of V. album differ, according to its host tree. However, little is known about the host-dependent phytochemical diversity in V. album . In this study, the metabolic profiles of V. album ssp. album from Malus domestica Bork., Quercus robur L., and Ulmus carpinifolia Gled were compared. Leaves, stems, and berries were collected in Switzerland, by the same procedure, in September 2016 and 2017. The methanolic extracts were analyzed by ultra-performance liquid chromatography, coupled to electrospray quadrupole time-of-flight mass spectrometry in positive ionization mode. The data were submitted to partial-least square discriminant analysis (PLS-DA) and the results showed that the V. album ssp. album samples were clustered into three groups, according to the three distinct host trees. Seven compounds, with high VIP scores (variable importance in projection), were responsible for this differentiation. The following four compounds were detected in both the harvest years: arginine, pipecolic acid or lysine, dimethoxycoumarin, and sinapyl alcohol, suggesting their use as host specific V. album biomarkers. The present work highlights the importance of standardized harvest and analytical procedures for the reproducibility of the chemical results of herbal materials.

Leishmaniasis is a neglected infectious disease caused by protozoan parasites from Leishmania genus species, affecting millions of people, in several countries. The current available treatment for cutaneous leishmaniasis (CL) has presented many side effects. In this way, micro- and nanotechnology are important processes, since they may be useful for release profile modulation of CL drugs improving their bioavailability. Amphotericin B (AmB) is a macrolide antibiotic used as a second-choice treatment. This study aimed the development of oil-water nanoemulsions (NEs) containing AmB for topical administration to treat CL. Furthermore, NEs were characterized by their droplet size, morphology, drug content, stability, in vitro release profile, and ex vivo skin permeation. In vitro anti-leishmanial activity using Leishmania amazonensis promastigotes was also evaluated. NEs containing AmB presented droplet size lower than 60 nm with a polydispersity index lower than 0.5. The best AmB-NEs were submitted to stability tests and these formulations presented excellent results after 365 days under refrigeration, confirming the maintenance of the drug content higher than 95%. AmB-NEs displayed slow and controlled AmB kinetic release and low skin permeation. These formulations presented lower cytotoxicity in comparison with free AmB and higher anti-leishmanial effect against L. amazonensis promastigotes. Therefore, the selected AmB-NE formulations, especially AmB-NE01, presented promising results as novel alternatives for CL treatment. Graphical abstract.

Fermented aqueous extracts of Viscum album L. are widely used for cancer treatment in complementary medicine. The high molecular weight compounds viscotoxins and lectins are considered to be the main active substances in the extracts. However, a vast number of small molecules (≤1500 Da) is also expected to be present, and few studies have investigated their identities. In this study, a comprehensive metabolome analysis of samples of fermented aqueous extracts of V. album from two host tree species ( Malus domestica and Pinus sylvestris ), both prepared by two pharmaceutical manufacturing processes, was performed by liquid chromatography-high resolution tandem mass spectrometry (LC-HRMS/MS). A total of 212 metabolites were putatively annotated, including primary metabolites (e.g., amino acids, organic acids, etc.) and secondary metabolites (mostly phenolic compounds). A clear separation between V. album samples according to the host tree species, but not due to manufacturing processes, was observed by principal component analysis. The biomarkers responsible for this discrimination were assessed by partial least squares-discriminant analysis. Because V. album extracts from different host trees have different clinical applications, the present work highlights the possibility of characterizing the metabolome for identification and traceability of V. album fermented aqueous extracts.

Background: Viscum album L. (Santalaceae), commonly known as mistletoe, is a hemiparasitic plant traditionally used in complementary cancer treatment. Its antitumor potential is mostly attributed to the presence of aqueous soluble metabolites; however, the use of ethanol as solvent also permits the extraction of pharmacological compounds with antitumor potential. The clinical efficacy of mistletoe therapy inspired the present work, which focuses on ethanolic extracts (V. album "mother tinctures", MT) prepared from different host trees., Methods: Samples from three European subspecies (album, austriacum, and abietis) were harvested, and five different V. album-MT strains were prepared. The following phytochemical analyses were performed: thin layer chromatography (TLC), high-performance liquid chromatography (HPLC) and liquid chromatography-high resolution mass spectrometry (LC-HRMS). The proliferation assay was performed with WST-1 after incubation of tumor (Yoshida and Molt-4) and fibroblast cell lines (NIH/3 T3) with different MT concentrations (0.5 to 0.05% v/v). The cell death mechanism was investigated by flow cytometry (FACS) using Annexin V-7AAD., Results: Chemical analyses of MT showed the presence of phenolic acids, flavonoids and lignans. The MT flavonoid and viscotoxin contents (mg/g fresh weight) were highest in Quercus robur (9.67 ± 0.85 mg/g) and Malus domestica (3.95 ± 0.58 mg/mg), respectively. The viscotoxin isoform proportions (% total) were also different among the VA subspecies with a higher content of A3 in V. album growing on Abies alba (60.57 ± 2.13). The phytochemical compounds as well as the viscotoxin contents are probably related to the antitumor effects of MT. The cell death mechanisms evaluated by colorimetric and FACS methodologies involved necrotic damage, which was host tree-, time- and dose- dependent, with different selectivity to tumor cells. Mother tincture from V. album ssp. abietis was the most effective at inducing in vitro cellular effects, even when incubated at the smallest concentration tested, probably because of the higher content of VT A3., Conclusion: Our results indicate the promising antitumor potential of Viscum album ethanolic extracts and the importance of botanical and phytochemical characterization for in vitro anti-proliferative effects.

Purpose: The present randomized controlled clinical trial evaluated the efficacy of homeopathic medicines of Melissa officinalis (MO), Phytolacca decandra (PD), and the combination of both in the treatment of possible sleep bruxism (SB) in children., Study Design: Patients (n = 52) (6.62 ± 1.79 years old) were selected based on the parents report of SB. The study comprised a crossover design that included 4 phases of 30-day treatment (Placebo; MO 12c; PD 12c; and MO 12c + PD 12c), with a wash-out period of 15 days between treatments., Methods: At baseline and after each phase, the Visual Analogic Scale (VAS) was used as the primary outcome measure to evaluate the influence of treatments on the reduction of SB. The following additional outcome measures were used: a children's sleep diary with parent's/guardian's perceptions of their children's sleep quality, the trait of anxiety scale (TAS) to identify changes in children's anxiety profile, and side effects reports. Data were analyzed by ANOVA with repeated measures followed by Post Hoc LSD test., Results: Significant reduction of SB was observed in VAS after the use of Placebo (-1.72 ± 0.29), MO (-2.36 ± 0.36), PD (-1.44 ± 0.28) and MO + PD (-2.21 ± 0.30) compared to baseline (4.91 ± 1.87). MO showed better results compared to PD (p = 0.018) and Placebo (p = 0.050), and similar result compared to MO+PD (p = 0.724). The sleep diary results and TAS results were not influenced by any of the treatments. No side effects were observed after treatments., Conclusion: MO showed promising results in the treatment of possible sleep bruxism in children, while the association of PD did not improve MO results., (Copyright © 2019. Published by Elsevier GmbH.)

Introduction:  According to the "silica hypothesis" formulated to explain homeopathy, the information of starting materials would be transferred to cells by silica nanoparticles detached from the glassware walls by serial dilution and agitation through epitaxy. We compared the biological activity, electrical current and silicon microparticle content (by means of scanning electron microscopy/energy-dispersive X-ray spectroscopy) of high dilutions (HDs) of arsenic prepared in plastic and glass vials to investigate the role of silica in their biological effects in vitro., Materials and Methods:  Co-cultures of macrophages and yeast ( Saccharomyces cerevisiae ) were treated with different HDs of arsenic prepared in plastic and glass vials. Macrophage morphology, phagocytosis index, nitric oxide (NO), and cytokine production were evaluated., Results:  Measurable amounts of silicon microparticles were detected only in the HDs prepared in glass vials, but ultra-centrifugation eliminated them. Specific and non-specific results were observed. Non-specific pro-inflammatory effects were seen in all dilutions prepared in plastic vials, including elevation of pro-inflammatory cytokines, NO and macrophage phagocytic index. Only the 200th centesimal dilution of arsenic produced specific decrease in interleukin-6 production in macrophages, and it was independent of the vial type or the presence of microparticles of silica in the medicine samples. The nature of the vials had an impact on the electric flow in the respective fluids., Conclusion:  The non-specific, pro-inflammatory effects might be attributed to organic residuals detached from the vials' plastic walls during manipulation. Instead, specific silica-independent effects of the homeopathic medicine can be attributed to the decrease of interleukin-6 after treatment with the 200th centesimal dilution of arsenic., Competing Interests: The authors declare there is no conflict of interest. Funding sponsors did not interfere with the design of the study, collection, analysis or interpretation of data, manuscript writing, or with the decision to publish the results. The initial project was selected for ABFH grant after ad hoc and blind evaluation of the proposal., (The Faculty of Homeopathy.)

Cancer is one of the biggest problems in public health worldwide. Plants have been shown important role in anticancer research. Viscum album L. (Santalaceae), commonly known as mistletoe, is a semi-parasitic plant that grows on different host trees. In complementary medicine, extracts from European mistletoe ( Viscum album L.) have been used in the treatment of cancer. The study was conducted to identify chemical composition and antitumor potential of Viscum album tinctures. Chemical analysis performed by high resolution chromatography equipped with high resolution mass spectrometer identified caffeic acid, chlorogenic acid, sakuranetin, isosakuranetin, syringenin 4-O-glucoside, syringenin 4-O-apiosyl-glucoside, alangilignoside C and ligalbumoside A compounds. Some of these compounds are probably responsible for the reduction of tumoral cellular growth in a dose-dependent manner. It was observed that melanoma murine cells (B16F10) were more sensitive to V. album tinctures than human leukaemic cells (K562), besides non-tumoral cells (MA-104) had a much lower cytotoxicity to them. Apoptotic-like cells were observed under light microscopy and were confirmed by a typical DNA fragmentation pattern. Additionally, flow cytometry results using Annexin-V/FITC permitted to quantify increased expression of early and late apoptotic markers on tumoral cells, confirming augmented Sub G0 population, which was probably associated with a consistent decrease in G1, and an increase in S or G2/M populations. Results indicate the chemical composition of V. album tinctures influences the mechanisms of in vitro tumoral cell death, suggesting a potential use in cancer pharmacotherapy research.

Background: A series of different experimental approaches was applied in Zincum metallicum (Zinc met.) samples and lactose controls. Experiments were designed to elucidate the effect of zinc trituration and dynamization on physicochemical properties of homeopathic formulations, using lactose as excipient., Methods: Zinc met. potencies (Zinc met 1-3c) were triturated and dynamized using lactose as excipient, according to Brazilian Homeopathic Pharmacopoeia. Lactose samples (LAC 1-3c) were also prepared following the same protocol and used as controls. The samples were analyzed structurally by Atomic Absorption Spectroscopy (AAS), X-ray Diffraction (XRD), Transmission Electron Microscopy (TEM) with Energy Dispersive X-ray Spectroscopy (EDX) and Scanning Electron Microscopy (SEM), and thermodynamically by Thermogravimetry (TG) and Differential Scanning Calorimetry (DSC)., Results: AAS analysis detected 97.0 % of zinc in the raw material, 0.75 % (Zinc met 1c) and 0.02% (Zinc met 2c). XRD analysis showed that inter-atomic crystalline spacing of lactose was not modified by dynamization. Amorphous and crystalline lactose spheres and particles, respectively, were observed by TEM in all samples, with mean size from 200 to 800 nm. EDX obtained with TEM identified zinc presence throughout the amorphous matter but individualized zinc particles were not observed. SEM images obtained from dynamized samples (LAC 1c and Zinc met 1c) with electron backscattering could not identify zinc metal grains. The dynamization process induced Derivatives of Thermal Gravimetric (DTg) peak modification, which was previously centered near 158°C to lactose, to a range from 140 to 170°C, suggesting the dynamization process modifies the temperature range of water aggregation. Thermal phenomena were analyzed and visualized by Analysis of Variance (ANOVA) and Principal Component Analysis (PCA) statistics. Both indicated that fusion enthalpy of dynamized samples (DynLAC 1-3c; DynZn 1-3c) increased 30.68 J/g in comparison to non-dynamized lactose (LAC; p < 0.05)., Conclusions: Our results suggested no structural changes due to the trituration and dynamization process. However, TG and DSC analyses permit the differentiation of dynamized and non-dynamized groups, suggesting the dynamization process induced a significant increase in the degradation heat. These results call for further calorimetric studies with other homeopathic dilutions and other methodologies, to better understand the dynamics of these systems., (Copyright © 2017 The Faculty of Homeopathy. Published by Elsevier Ltd. All rights reserved.)

The prevalence of Th1/Th2 response, spleen changes and megakaryocytes were investigated in BALB/c mice (n=138) infected with Leishmania infantum, and treated with Leishmania infantum 30× (10 -30 ) biotherapy - BioLi30×. We performed controlled experiments using 8-to-12-week-old mice, infected with 5×10 7 L. infantum promastigotes, divided into eight groups: G1 (healthy), G2 (infected with L. infantum), G3 (BioLi30× pre-treated), G4 (BioLi30× pre/post-treated), G5 (BioLi30× post-treated), G6 (Water 30× post-treated), G7 (Antimonium crudum 30× post-treated) and G8 (Glucantime® post-treated). G3-G7 groups were orally treated with their respective drugs diluted in filtered water (1:10), and G8 received Glucantime® (0.6mg/100µl of PBS), intraperitoneally. Spleen fragments were submitted to double blind histopathological evaluation and the number of megakaryocytes was counted. Besides, animals' serum was measured after 49days of infection, and cytokines (IFN-γ, IL-4, IL-10, IL-12), as well as the Th1/Th2 correlation (IFN-γ/IL-4 and IFN-γ/IL-10), were analyzed. Spleen histological parameters were classified as: healthy appearance (G1); discreet (G3-G7), moderate (G2) and moderate to severe (G8) white pulp hyperplasia; proliferation of megakaryocytes (G2-G8), and intense disruption (G2-G8). All groups, except for G7, showed higher percentages of megakaryocytes per field ranging from 87% to 15%, when compared to healthy animals (G1). Th1 predominance in IFN-γ/IL-4 ratio (comparing to G2) was detected in G4, G5, G6 and G7. Finally, pre/post (BioLi30x) and post-treatment (Antimonium crudum 30x) presented reduction of megakaryocytes/spleen changes due to immunomodulation animal process, controlling the infection process, probably by the Th1 cytokine predominance., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Available treatments against human fungal pathogens present high levels of resistance, motivating the development of new antifungal therapies. In this context, the present work aimed to analyze direct electric current (DC) antifungal action, using an in vitro apparatus equipped with platinum electrodes. Candida albicans yeast cells were submitted to three distinct conditions of DC treatment (anodic flow-AF; electroionic flow-EIF; and cathodic flow-CF), as well as different charges, ranging from 0.03 to 2.40 C. Our results indicated C. albicans presented distinct sensibility depending on the DC intensity and polarity applied. Both the colony-forming unit assay and the cytometry flow with propidium iodide indicated a drastic reduction on cellular viability after AF treatment with 0.15 C, while CF- and EIF-treated cells stayed alive when DC doses were increased up to 2.40 C. Additionally, transmission electron microscopy revealed important ultrastructural alterations in AF-treated yeasts, including cell structure disorganization, ruptures in plasmatic membrane, and cytoplasmic rarefaction. This work emphasizes the importance of physical parameters (polarity and doses) in cellular damage, and brings new evidence for using electrotherapy to treat C. albicans pathology process. Bioelectromagnetics. 38:95-108, 2017. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

Electrochemical therapy (EChT) entails treatment of solid tumors with direct electric current (DC). This work evaluated the specific effects of anodic flow generated by DC on biochemical and metabolic features of the A549 human lung cancer cell line. Apoptosis was evaluated on the basis of caspase-3 activity and mitochondrial transmembrane potential dissipation. Cell morphology was analyzed using transmission electron microscopy, and lipid droplets were studied through morphometric analysis and X-ray qualitative elemental microanalysis. High-resolution respirometry was used to assess mitochondrial respiratory parameters. Results indicated A549 viability decreased in a dose-dependent manner with a prominent drop between 18 and 24h after treatment (p<0.001), together with a two-fold increase in caspase-3 activity. AF-treatment induced a significantly increase (p<0.01) in the cell number with disrupted mitochondrial transmembrane potential. Furthermore, treated cells demonstrated important ultrastructural mitochondria damage and a three-fold increase in the cytoplasmic lipid bodies' number, quantified by morphometrical analyses. Conversely, 24h after treatment, the cells presented a two-fold increase of residual oxygen consumption, accounting for 45.3% of basal oxygen consumption. These results show remarkable alterations promoted by anodic flow on human lung cancer cells which are possibly involved with the antitumoral effects of EChT., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Background: Influenza affects thousands of people worldwide every year, motivating the development of new therapies. In this work, the effects of two homeopathic preparations (influenza biotherapies and thymulin) were chosen following two different rationales: isotherapy and endo-isotherapy models. The homeopathic effects were evaluated individually considering the inflammatory and behavioral responses against influenza virus antigen were studied in BALB/c mice., Methods: Male adult mice were treated orally and blindly for 21 days with highly diluted influenza virus or with thymulin, and were divided in two sets of experiments. The first series of experiments aimed to describe their behavior, using an open field (OF) device. In the second series, mice were challenged subcutaneously with influenza hemagglutinin antigen (7 μg/200 μl) at day 21. At day 42, behavior and inflammation response were evaluated., Results: No behavioral changes were seen in OF tests at any time point after treatments. Flow cytometry and morphometry revealed significant changes in T and B cell balance after influenza antigen challenge, varying according to treatment., Conclusion: The results show that both homeopathic treatments induced subtle changes in acquired immune anti-viral response regulation. A deeper understanding of the mechanism could elucidate their possible use in influenza epidemiological situations., (Copyright © 2016 The Faculty of Homeopathy. Published by Elsevier Ltd. All rights reserved.)

The Franz cells permeation assay has been performed for over 25 years. However, the advent of nanotechnology created a whole new world, especially with regard to topical products. In this new global scenario an increasing number of nanostructure-based delivery systems (NDSs) have emerged and a global warning relating to the safety of these NDSs is arising. This work studied the efficacy of the Franz cells assay, comparing it with the radiolabeling biodistribution test. For this purpose a formulation of sunscreen based on an NDS was developed and characterized. The results demonstrated both that the NDS did not present in vitro cytotoxicity and that the radiolabeling biodistribution test is more precise for the evaluation of NDS cosmetics than the Franz cells assay, since it detected the permeation of the NDS at a picogram order. Due to this fact, and considering all the concerns related to NDSs and nanoparticles in general, more precise methods must be used in order to guarantee the safe use of these new classes of products.

Objectives: The aim of this study was to evaluate the effect of SLPI on the growth and biological processes of Candida albicans., Methods: Two C. albicans strains were used in this study, a clinical isolate resistant to fluconazole (PRI) and a reference strain ATCC 24433. The minimal inhibitory concentration (MIC) was determined according to the CLSI methodology. The influence of SLPI on secreted serine proteinase activities (SSP) was measured by the cleavage of specific substrate, and surface hydrophobicity was determined by the aqueous-hydrocarbon biphasic separation method. Flow cytometry was performed to investigate receptors for SLPI and variations in the cell wall mannoprotein expression. Interaction between yeast and epithelium was assessed using the MA-104 cells lineage. Ultrastructure was analyzed by transmission electron microscopy (TEM)., Results: MIC values were calculated as 18 and 18.9μM for the PRI and ATCC 24433, respectively. SSP activity was reduced by 48.8% by 18μM of SLPI and cell surface hydrophobicity increased by 11.1%. Flow cytometry suggest the existence of SLPI binding sites on the surface of the yeast. Results showed a reduction in the expression of mannoproteins in 20.8% by the cells treated with 80μM of SLPI, and 18μM reduced the adhesion of yeasts to mammalian cells in 60.1%. TEM revealed ultrastructural changes in cells treated with 80μM of SLPI, such as the presence of membrane-like structures within the cytoplasm., Conclusions: SLPI exerts a significant influence on C. albicans viability and biological processes. Considering its constitutive and physiologic features, SLPI may become a promising tool for the development of new methodologies for the treatment and control of candidiasis., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Electrochemical treatment is an alternative modality for tumor treatment based on the application of a low intensity direct electric current to the tumor tissue through two or more platinum electrodes placed within the tumor zone or in the surrounding areas. This treatment is noted for its great effectiveness, minimal invasiveness and local effect. Several studies have been conducted worldwide to evaluate the antitumoral effect of this therapy. In all these studies a variety of biochemical and physiological responses of tumors to the applied treatment have been obtained. By this reason, researchers have suggested various mechanisms to explain how direct electric current destroys tumor cells. Although, it is generally accepted this treatment induces electrolysis, electroosmosis and electroporation in tumoral tissues. However, action mechanism of this alternative modality on the tumor tissue is not well understood. Although the principle of Electrochemical treatment is simple, a standardized method is not yet available. The mechanism by which Electrochemical treatment affects tumor growth and survival may represent more complex process. The present work analyzes the latest and most important research done on the electrochemical treatment of tumors. We conclude with our point of view about the destruction mechanism features of this alternative therapy. Also, we suggest some mechanisms and strategies from the thermodynamic point of view for this therapy. In the area of Electrochemical treatment of cancer this tool has been exploited very little and much work remains to be done. Electrochemical treatment constitutes a good therapeutic option for patients that have failed the conventional oncology methods.

Background: Influenza viruses cause highly contagious acute respiratory illnesses with significant mortality, especially among young children, elderly people, and individuals with serious medical conditions. This encourages the development of new treatments for human flu. Biotherapies are diluted solutions prepared from biological products compounded following homeopathic procedures., Objectives: To develop a biotherapy prepared from the infectious influenza A virus (A/Aichi/2/68 H3N2) and to verify its in vitro response., Methods: The ultradiluted influenza virus solution was prepared in the homeopathic dilution 30dH, it was termed Influenzinum RC. The cellular alterations induced by this preparation were analyzed by optical and electron microscopy, MTT and neutral red assays. Glycolytic metabolism (PFK-1) was studied by spectrophotometric assay. Additionally, the production of tumor necrosis factor-α (TNF-α) by J774.G8 macrophage cells was quantified by ELISA before and after infection with H3N2 influenza virus and treatment., Results: Influenzinum RC did not cause cytotoxic effects but induced morphological alterations in Madin-Darby canine kidney (MDCK) cells. After 30 days, a significant increase (p < 0.05) in mitosis rate was detected compared to control. MDCK mitochondrial activity was changed after treatment for 10 and 30 days. Treatment significantly diminished (p < 0.05) PFK-1 activity. TNF-α in biotherapy-stimulated J774.G8 macrophages indicated a significant (p < 0.05) increase in this cytokine when the cell supernatant was analyzed., Conclusion: Influenzinum RC altered cellular and biochemical features of MDCK and J774G8 cells., (Copyright © 2012 The Faculty of Homeopathy. Published by Elsevier Ltd. All rights reserved.)

The literature shows that the effects of direct electric currents on biological material are numerous, including bactericidal, fungicidal, parasiticidal, and anti-tumoral, among others. Non-pathogenic trypanosomatids, such as Herpetomonas samuelpessoai, have emerged as important models for the study of basic biological processes performed by a eukaryotic cell. The present study reports a dose-dependent anti-protozoan effect of direct electric treatment with both cathodic and anodic current flows on H. samuelpessoai cells. The damaging effects can be attributable to the electrolysis products generated during electric stimulation. The pH of the cell suspension was progressively augmented from 7.4 to 10.5 after the cathodic treatment. In contrast, the anodic treatment caused a pH decrease varying from 7.4 to 6.5. Transmission electron microscopy analyses revealed profound alterations in vital cellular structures (e.g., mitochondrion, kinetoplast, flagellum, flagellar pocket, nucleus, and plasma membrane) after exposure to both cathodic and anodic current flows. Specifically, cathodic current flow treatment induced the appearance of autophagic-like structures on parasite cells, while those submitted to an anodic current flow presented marked disorganization of plasma membrane and necrotic appearance. However, parasites treated in the intermediary chamber (without contact with the electrodes) did not present significant changes in viability or morphology, and no pH variation was detected in this system. The use of H. samuelpessoai as a biological model and the direct electric current experimental approach used in our study provide important information for understanding the mechanisms involved in the cytotoxic effects of this physical agent., (Copyright © 2011 Wiley Periodicals, Inc.)

Staphylococcus lugdunensis is an unusually virulent coagulase-negative species, which causes serious infection similar to S. aureus. We evaluated the expression of virulence factors such as S. lugdunensis synergistic haemolysin (SLUSH), fibrinogen-binding protein (Fbl), biofilm production and biofilm-production-related genes in 23 S. lugdunensis clinical isolates and one type strain that had been previously characterized for their genotypes. In addition, the biofilm composition and the ability of isolates to adhere to and invade human epithelial lung cells were also investigated. The PCR method used detected the presence of slush and intercellular adhesin (ica) virulence genes in all isolates. All isolates produced the Fbl protein and, with the exception of the type strain, all isolates produced the SLUSH haemolysin. Fourteen (60.9 %) isolates produced biofilms. The detachment assay, using sodium metaperiodate or proteolytic enzymes to analyse the biofilm composition, showed protein-mediated biofilms in two representative isolates, one for each colony type (rough and smooth). All strongly biofilm-producing isolates, including three with rough colony morphology, had the same prevalent PFGE pattern. However, among the representative strains tested, only the S. lugdunensis isolate that formed rough colonies was able to adhere to and invade A549 cell monolayers in the same quantities as those observed with S. aureus isolates (P = 1.000). No significant adhesion or invasion was observed for the other isolates in comparison with the S. aureus isolate, independent of biofilm production or clonality. Our results could explain the incredible ability of this pathogen to cause infections that are as aggressive as S. aureus. In addition, the ability of S. lugdunensis to adhere to and invade eukaryotic cells was also noticed for isolates with rough colony morphology, reinforcing the increased virulence in this species.

Inhibition of tumor growth induced by treatment with direct electric current (DC) has been reported in several models. One of the mechanisms responsible for the antitumoral activity of DC is the generation of oxidative species, known as chloramines. With the aim of increasing chloramine production in the electrolytic medium and optimizing the antitumoral effects of DC, poly(ɛ-caprolactone) (PCL) nanoparticles (NPs) loaded with the amino acid tyrosine were obtained. The physical-chemical characterization showed that the NPs presented size in nanometric range and monomodal distribution. A slightly negative electrokinetic potential was also found in both blank NPs and L-tyrosine-loaded PCL NPs. The yield of the loading process was approximately 50%. Within 3 h of dissolution assay, a burst release of about 80% L-tyrosine was obtained. The in vitro cytotoxicity of DC was significantly increased when associated with L-tyrosine-loaded NPs, using a murine multidrug-resistant melanoma cell line model. This study showed that the use of the combination of nanotechnology and DC has a promising antineoplastic potential and opens a new perspective in cancer therapy.