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Reports from popular medicine usually act as a basis for the development of new drugs from natural compounds with therapeutic actions for serious diseases and prevalence such as cancer. Bromelia antiacantha Bertol. is a species of the Bromeliaceae family, considered an unconventional food plant, found in the south and midwest regions of Brazil. Despite the high nutritional content and pharmacological potential of its fruits, few scientific studies report its biological actions. Thus, this study evaluates the phytochemical profile of aqueous and ethanol extracts obtained from B. antiacantha fruits, as well as their possible antioxidant, antitumor, and cytotoxic activities. The aqueous extract exhibited phenolic compounds and flavonoids, while ethanol extracts indicated the presence of flavonoids and coumarin in their composition, regardless of the region of collection. The ethanolic extract demonstrated a more promising antioxidant effect than the aqueous extract and also induced a significant inhibition in the viability of human cervical cancer cells of the SiHa strain. In addition, treatment with both extracts did not alter the viability of non-tumor cells of the immortalized human keratinocyte lineage (HaCaT). These results bring new data about extracts obtained from a native plant, edible and traditionally used in popular medicine, opening new perspectives for its possible therapeutic application.

Relatos da medicina popular costumam atuar como referencial para o desenvolvimento de novos fármacos a partir de moléculas naturais com ações terapêuticas para doenças de alta gravidade e prevalência como o câncer. Bromelia antiacantha Bertol. é uma espécie da família Bromeliaceae, considerada uma planta alimentícia não convencional (PANC), encontrada nas regiões sul e centro-oeste do Brasil. Apesar do alto teor nutritivo e potencial farmacológico de seus frutos, poucos estudos científicos relatam suas ações biológicas. Desta forma, este estudo avalia o perfil fitoquímico de extratos aquoso e etanólico obtidos de frutos de B. antiacantha, bem como a sua possível ação antioxidante, antitumoral e citotóxica. O extrato aquoso apresentou compostos fenólicos e flavonoides, enquanto os extratos etanólicos apontam a presença de flavonóides e cumarina em sua composição, independente da região de coleta. O extrato etanólico demonstrou efeito antioxidante mais promissor do que o extrato aquoso e também induziu uma inibição significativa na viabilidade de células humanas de câncer cervical da linhagem SiHa. Além disso, o tratamento com ambos extratos não alterou a viabilidade de células não tumorais da linhagem de queratinócitos humanos imortalizados (HaCaT). Estes dados trazem novas informações sobre extratos obtidos de uma espécie vegetal nativa, comestível e já utilizada tradicionalmente, mas abrindo novas perspectivas quanto a possíveis aplicações terapêuticas.

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Abstract Reports from popular medicine usually act as a basis for the development of new drugs from natural compounds with therapeutic actions for serious diseases and prevalence such as cancer. Bromelia antiacantha Bertol. is a species of the Bromeliaceae family, considered an unconventional food plant, found in the south and midwest regions of Brazil. Despite the high nutritional content and pharmacological potential of its fruits, few scientific studies report its biological actions. Thus, this study evaluates the phytochemical profile of aqueous and ethanol extracts obtained from B. antiacantha fruits, as well as their possible antioxidant, antitumor, and cytotoxic activities. The aqueous extract exhibited phenolic compounds and flavonoids, while ethanol extracts indicated the presence of flavonoids and coumarin in their composition, regardless of the region of collection. The ethanolic extract demonstrated a more promising antioxidant effect than the aqueous extract and also induced a significant inhibition in the viability of human cervical cancer cells of the SiHa strain. In addition, treatment with both extracts did not alter the viability of non-tumor cells of the immortalized human keratinocyte lineage (HaCaT). These results bring new data about extracts obtained from a native plant, edible and traditionally used in popular medicine, opening new perspectives for its possible therapeutic application.

Resumo Relatos da medicina popular costumam atuar como referencial para o desenvolvimento de novos fármacos a partir de moléculas naturais com ações terapêuticas para doenças de alta gravidade e prevalência como o câncer. Bromelia antiacantha Bertol. é uma espécie da família Bromeliaceae, considerada uma planta alimentícia não convencional (PANC), encontrada nas regiões sul e centro-oeste do Brasil. Apesar do alto teor nutritivo e potencial farmacológico de seus frutos, poucos estudos científicos relatam suas ações biológicas. Desta forma, este estudo avalia o perfil fitoquímico de extratos aquoso e etanólico obtidos de frutos de B. antiacantha, bem como a sua possível ação antioxidante, antitumoral e citotóxica. O extrato aquoso apresentou compostos fenólicos e flavonoides, enquanto os extratos etanólicos apontam a presença de flavonóides e cumarina em sua composição, independente da região de coleta. O extrato etanólico demonstrou efeito antioxidante mais promissor do que o extrato aquoso e também induziu uma inibição significativa na viabilidade de células humanas de câncer cervical da linhagem SiHa. Além disso, o tratamento com ambos extratos não alterou a viabilidade de células não tumorais da linhagem de queratinócitos humanos imortalizados (HaCaT). Estes dados trazem novas informações sobre extratos obtidos de uma espécie vegetal nativa, comestível e já utilizada tradicionalmente, mas abrindo novas perspectivas quanto a possíveis aplicações terapêuticas.

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Cervical cancer (CC) is a serious global health issue, and it is well-known that HPV infection is the main etiological factor that triggers carcinogenesis. In cancer, chemokine ligands and receptors are involved in tumor cell growth, metastasis, leukocyte infiltration, and angiogenesis; however, information on the role played by E6/E7 of HPV16/18 in the modulation of chemokines is very limited. Therefore, this study aimed to determine whether chemokines are differentially expressed in CC-derived cell lines; if E6/E7 oncoproteins from HPV16 and 18 are capable of mediating chemokine expression, what is the expression profile of chemokines in tissues derived from CC and what is their impact on the overall survival of patients with this pathology? For this purpose, RNA sequencing and real-time PCR were performed on SiHa, HeLa, and C33A tumorigenic cell lines, on the non-tumorigenic HaCaT cells, and the E6/E7 HPV-transduced HaCaT cell models. Furthermore, chemokine expression and survival analysis were executed on 304 CC and 22 normal tissue samples from The Cancer Genome Atlas (TCGA) repository. The results demonstrate that CXCL1, CXCL2, CXCL3, and CXCL8 are regulated by E6/E7 of HPV16 and 18, are overexpressed in CC biopsies, and that their higher expression is related to a worse prognostic survival.

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The accumulated dental biofilm can be a source of oral bacteria that are aspirated into the lower respiratory tract causing ventilator-associated pneumonia in hospitalized patients. The aim of this study was to evaluate the synergistic antibiofilm action of the produced and phytochemically characterized extracts of Cinnamomum verum and Brazilian green propolis (BGP) hydroethanolic extracts against multidrug-resistant clinical strains of Acinetobacter baumannii and Pseudomonas aeruginosa, in addition to their biocompatibility on human keratinocyte cell lines (HaCaT). For this, High-performance liquid chromatography analysis of the plant extracts was performed; then the minimum inhibitory and minimum bactericidal concentrations of the extracts were determined; and antibiofilm activity was evaluated with MTT assay to prevent biofilm formation and to reduce the mature biofilms. The cytotoxicity of the extracts was verified using the MTT colorimetric test, evaluating the cellular enzymatic activity. The data were analyzed with one-way ANOVA and Tukey's tests as well as Kruskal-Wallis and Dunn's tests, considering a significance level of 5%. It was possible to identify the cinnamic aldehyde in C. verum and p-coumaric, caffeic, and caffeoylquinic acids as well as flavonoids such as kaempferol and kaempferide and Artepillin-C in BGP. The combined extracts were effective in preventing biofilm formation and reducing the mature biofilms of A. baumannii and P. aeruginosa. Moreover, both extracts were biocompatible in different concentrations. Therefore, C. verum and BGP hydroethanolic extracts have bactericidal and antibiofilm action against multidrug resistant strains of A. baumannii and P. aeruginosa. In addition, the combined extracts were capable of expressively inhibiting the formation of A. baumannii and P. aeruginosa biofilms (prophylactic effect) acting similarly to 0.12% chlorhexidine gluconate.

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Acinetobacter baumannii is a multi-drug resistant microorganism. The objective of this study was to evaluate the antimicrobial and antibiofilm action of the pomegranate natural extract against eight strains of multidrug resistant Acinetobacter baumanii and to assess the extract cytotoxicity in a culture of Human Keratinocytes (HaCat). Broth microdilution method was used to determine the minimum inhibitory and minimum microbicidal concentrations of the extracts. The extract antibiofilm action was analysed by the MTT colorimetric test. The cytotoxicity evaluation was performed by the MTT colorimetric test, which analysed the mitochondrial cellular action, after contact of the extract for 5 min. The results were statistically analysed by ANOVA and Tukey test with a significance level α≤ 0.05. Punica granatum L. (pomegranate) extract had antimicrobial action on all the 8 clinical strains of Acinetobacter baumannii evaluated. The extract showed a significant reduction in metabolic action in biofilm for all the strains, with results statistically different from growth control (p≤0.05%). P. granatum extract applied for 5 min on human keratinocytes (HaCat) promoted cell viability in all the tested concentrations. The pomegranate extract is effective in reducing the multidrug-resistant clinical strains of Acinetobacter baumanni and is biocompatible. (AU)

Acinetobacter baumannii é um microrganismo multirresistente. O objetivo deste estudo foi avaliar a ação antimicrobiana e antibiofilme do extrato natural de romã contra oito cepas de A. baumanii multirresistente e avaliar a citotoxicidade do extrato em uma cultura de queratinócitos humanos (HaCat). O método de microdiluição em caldo foi utilizado para determinar as concentrações inibitórias mínimas e microbicidas mínimas dos extratos. A ação antibiofilme do extrato foi analisada pelo teste colorimétrico MTT. A avaliação da citotoxicidade foi realizada pelo teste colorimétrico MTT, que analisou a ação celular mitocondrial, após contato do extrato por 5 min. Os resultados foram analisados ​​estatisticamente por ANOVA e teste de Tukey com nível de significância α≤ 0,05. O extrato de Punica granatum L. (romã) apresentou ação antimicrobiana em todas as 8 cepas clínicas avaliadas de A. baumannii. O extrato apresentou redução significativa na ação metabólica no biofilme para todas as linhagens, com resultados estatisticamente diferentes do controle de crescimento (p≤0,05%). O extrato de P. granatum aplicado por 5 min em queratinócitos humanos (HaCat) promoveu viabilidade celular em todas as concentrações testadas. O extrato de romã é eficaz na redução das cepas clínicas multirresistentes de Acinetobacter baumanni e é biocompatível. (AU)

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The pathogenesis of atopic dermatitis (AD) and psoriasis (Ps) overlaps, particularly the activation of the immune response and tissue damage. Here, we evaluated galectin (Gal)-1 and Gal-3 levels, which are beta-galactoside-binding proteins with immunomodulatory functions and examined their effects on human keratinocytes stimulated with either interleukin (IL)-4 or IL-17A. Skin biopsies from AD, Ps, and control patients were evaluated using histological and immunohistochemical analyses. Six studies containing publicly available transcriptome data were individually analyzed using the GEO2R tool to detect Gal-1 and Gal-3 mRNA levels. In vitro, IL-4- or IL-17A-stimulated keratinocytes were treated with or without Gal-1 or Gal-3 to evaluate cytokine release and migration. Our findings showed different patterns of expression for Gal-1 and Gal-3 in AD and Ps skins. Densitometric analysis in skin samples showed a marked increase in the protein Gal-1 levels in Ps epidermis and in both AD and Ps dermis compared to controls. Protein and mRNA Gal-3 levels were downregulated in AD and Ps lesional skin compared with the control samples. In vitro, both galectins addition abrogated the release of IL-8 and RANTES in IL-17-stimulated keratinocytes after 24 h, whereas IL-6 release was downregulated by Gal-3 and Gal-1 in IL-4- and IL-17-stimulated cells, respectively. Administration of both galectins also increased the rate of keratinocyte migration under IL-4 or IL-17 stimulation conditions compared with untreated cells. Altogether, the immunoregulatory and migration effects of Gal-1 and Gal-3 on keratinocytes under inflammatory microenvironment make them interesting targets for future therapies in cutaneous diseases.

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Abstract: The present study aimed to characterize the chemical elements and cytotoxicity of Carnoy's solution (CS) by comparing two different trademarked products (one Brazilian [NCS] and another imported [ICS]) using inductively coupled plasma mass spectrometry (ICP-MS) and human keratinocyte (HaCaT) cultures. For performing ICP-MS, the solutions were diluted according to calibration curves, and the chemical elements were analyzed with a spectrometer. HaCaT cells were exposed to CS concentrations ranging from 0.10% to 20% for 3 or 5 min. Cell viability was evaluated immediately (T0), 24 h (T1), and 7 days (T2) after exposure to CS using 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyl-tetrazolium bromide (MTT) reduction assay. Data were analyzed using a t-test for ICP-MS and analysis of variance followed by Tukey's post-hoc test for MTT assay, both considering statistical significance at p<0.05. ICP-MS results revealed that ICS presented significantly lower concentrations of 12 chemical elements than NCS. The results of MTT assay revealed that at T0, ICS was more cytotoxic than NCS regardless of the time of exposure (p < 0.05). At T1, the only difference between the groups was at a concentration of 0.10% after 5 min of exposure. At T2, at a concentration of 0.5%, ICS resulted in a significant reduction in cell viability compared to NCS (p < 0.05). Thus, the results showed that ICS was more cytotoxic than NCS. Collectively, our findings suggest that the individual compositions of different CS formulations should be investigated.

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Tattoo inks represent a growing market in the world economy, but this growth is associated with an increase in reports of adverse effects caused by the use of this product. In this study, four commercial tattoo inks (blue, green, red and black) were studied to characterize the composition and particle size and identify possible in vivo and in vitro toxicological effects on Daphnia magna and HaCaT cells, respectively. Compositional analysis confirmed the functional groups in the vehicles and organic pigments. The presence of nanoparticles was confirmed by image analysis. The toxicological evaluation indicated distinct results for blue and green inks for the parameters tested, despite the presence of similar levels of metals. The red ink, followed by the green, presented the highest toxicity, which may be related to pigments containing azo compounds and not to the metal fraction. Black ink was found to be the safest toxicologically. This paper provides an overview of the composition of tattoo inks and their toxicological effects in epidermal cells and in the environment.

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Although the effects of low-intensity pulsed ultrasound (LIPUS) on diverse cell types have been fully studied, the functional role of LIPUS in keratinocytes remains poorly understood. This study aimed to investigate the effects of LIPUS on proliferation and migration of HaCaT cells as well as the regulatory mechanisms associated with signaling pathways. Human HaCaT cells were exposed or not to LIPUS, and cell proliferation and migration were measured by BrdU incorporation assay and Transwell assay, respectively. Expression of proteins associated with proliferation and migration was evaluated by western blot analysis. Expression of key kinases in the PI3K/AKT and JNK pathways was also evaluated by western blot analysis. Effects of LIPUS on the PI3K/AKT and JNK pathways, and whether LIPUS affected HaCaT cells via these two pathways were finally explored. When the parameter of LIPUS (number of cycles) was set at 300, cell viability was the highest after LIPUS stimulation. We then found that the percentage of BrdU positive cells was enhanced by LIPUS, along with up-regulation of cyclinD1, CDK6, CDK4, and VEGF. LIPUS promoted migration, as well as up-regulation of MMP-2 and MMP-9. Phosphorylation levels of key kinases in the PI3K/AKT and JNK pathways were increased by LIPUS. Inhibition of either PI3K/AKT pathway or JNK pathway attenuated effects of LIPUS on HaCaT cells, and co-inhibition of these two pathways showed augmented effects. LIPUS promoted proliferation and migration of HaCaT cells through activating the PI3K/AKT and JNK pathways.

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BACKGROUND: Cervical cancer is the fourth cause of death worldwide by cancer in women and is a disease associated to persistent infection with human papillomavirus (HPV), particularly from two high-risk types HPV16 and 18. The virus initiates its replicative cycle infecting cells located in the basal layer of the epithelium, where a small population of epithelial stem cells is located performing important functions of renewal and maintenance of the tissue. Viral E2 gene is one of the first expressed after infection and plays relevant roles in the replicative cycle of the virus, modifying fundamental processes in the infected cells. Thus, the aim of the present study was to demonstrate the presence of hierarchic subpopulations in HaCaT cell line and evaluate the effect of HPV16-E2 expression, on their biological processes. METHODS: HaCaT-HPV16-E2 cells were generated by transduction of HaCaT cell line with a lentiviral vector. The α6-integrin-CD71 expression profile was established by immunostaining and flow cytometric analysis. After sorting, cell subpopulations were analyzed in biological assays for self-renewal, clonogenicity and expression of stemness factors (RT-qPCR). RESULTS: We identified in HaCaT cell line three different subpopulations that correspond to early differentiated cells (α6-integrindim), transitory amplifying cells (α6-integrinbri/CD71bri) and progenitor cells (α6-integrinbri/CD71dim). The last subpopulation showed stem cell characteristics, such as self-renewal ability, clonogenicity and expression of the well-known stem cell factors SOX2, OCT4 and NANOG, suggesting they are stem-like cells. Interestingly, the expression of HPV16-E2 in HaCaT cells changed its α6-integrin-CD71 immunophenotype modifying the relative abundance of the cell subpopulations, reducing significantly the percentage of α6-integrinbri/CD71dim cells. Moreover, the expression of the stem cell markers was also modified, increasing the expression of SOX2 and NANOG, but decreasing notably the expression of OCT4. CONCLUSIONS: Our data demonstrated the presence of a small subpopulation with epithelial "progenitor cells" characteristics in the HaCaT cell line, and that HPV16-E2 expression on these cells induces early differentiation.

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