New reports on secretory structures of vegetative and floral organs of Hypericum elodes (Hypericaceae)

Hypericum elodes L. (Hypericaceae), commonly known as marsh St. John’s wort, is one of the fourteen species found spontaneously in Portugal and it is endemic of Europe. It occurs in acidic waterlogged grounds in the Norwest of Portugal and in the Azores islands, Pico and São Miguel [1]. In the last decades intense phytochemical and pharmacological research have been performed in Hypericum species, namely in H. perforatum, the most studied species of the genus and traditionally used as a medicinal plant. Its bioactive secondary metabolites, naphtodianthrones (e.g. hypericin and pseudohypericin), phloroglucinols (e.g. hyperforin), bioflavonoids and xanthones have been widely studied for their anti-depressant, anti-microbial, anti-viral and anti-proliferative properties [2]. Despite the abundant phytochemical reports available in Hypericum species, the morpho-anatomical studies are scarce and fragmented [3-6]. The present study, included in an ongoing project on Hypericum glands, aims to provide new data on the morphology and anatomy of the secretory structures of H. elodes. Although these glands were previously studied in specimens grown in Italy [5], in the current study we describe in detail their morphology, distribution and histochemistry on the vegetative and floral organs. Small branches of H. elodes were collected from populations occurring in Portugal and samples of leaves and flowers, at different stages of development, were fixed with glutaraldehyde and prepared for scanning electron microscopy or embedding in Leica Historesin®, following standard methods. Additionally, the main classes of compounds present in the secretion were histochemically characterized

Secretory structures on the flowers of Hypericum Pubescens and H. Tomentosum

Hypericum L. (Hypericaceae) is a genus represented by ca. 484 species, shrubs, perennial and annual herbs, growing widely in warm-temperate areas in Europe, West Asia and America [1]. H. perforatum (St. John’s wort), the most representative member of the genus has been used in folk medicine since Antiquity, and nowadays other Hypericum species have been incorporated in traditional medicine systems of several countries around the world. It is now well known that hypericin, pseudohypericin and hyperforin are the main compounds responsible for their therapeutic properties [2]. Although a large number of studies deal with the phytochemical and pharmacological characteristics of Hypericum species, the secretory structures that produce the bioactive compounds have only been examined in detail in a few species [3, 4, 5, 6]. Within the framework of a wider project on Hypericum glands, we have undertaken cytological studies on the flowers of H. pubescens and H. tomentosum. Here we describe the structure, development and distribution pattern of the glands present in these two Hypericum species. Flowers at different stages of development were collected from natural populations of H. pubescens and H. tomentosum, occurring in Portugal. Samples fixed with glutaraldehyde were prepared for scanning electron microscopy or embedding in Leica Historesin® for anatomy, following standard methods

Nanoformulation of seaweed eisenia bicyclis in albumin nanoparticles targeting cardiovascular diseases: In vitro and in vivo evaluation

Natural products, especially those derived from seaweeds, are starting to be seen as effective against various diseases, such as cardiovascular diseases (CVDs). This study aimed to design a novel oral formulation of bovine albumin serum nanoparticles (BSA NPs) loaded with an extract of Eisenia bicyclis and to validate its beneficial health effects, particularly targeting hypercholesterolemia and CVD prevention. Small and well-defined BSA NPs loaded with Eisenia bicyclis extract were successfully prepared exhibiting high encapsulation efficiency. Antioxidant activity and cholesterol biosynthesis enzyme 3-hydroxy-3 methylutaryl coenzyme A reductase (HMGR) inhibition, as well as reduction of cholesterol permeation in intestinal lining model cells, were assessed for the extract both in free and nanoformulated forms. The nanoformulation was more efficient than the free extract, particularly in terms of HMGR inhibition and cholesterol permeation reduction. In vitro cytotoxicity and in vivo assays in Wistar rats were performed to evaluate its safety and overall effects on metabolism. The results demonstrated that the Eisenia bicyclis extract and BSA NPs were not cytotoxic against human intestinal Caco-2 and liver HepG2 cells and were also safe after oral administration in the rat model. In addition, an innovative approach was adopted to compare the metabolomic profile of the serum from the animals involved in the in vivo assay, which showed the extract and nanoformulation's impact on CVD-associated key metabolites. Altogether, these preliminary results revealed that the seaweed extract and the nanoformulation may constitute an alternative natural dosage form which is safe and simple to produce, capable of reducing cholesterol levels, and consequently helpful in preventing hypercholesterolemia, the main risk factor of CVDs.info:eu-repo/semantics/publishedVersio

Inhibition of HMG-CoA reductase activity and cholesterol permeation through Caco-2 cells by caffeoylquinic acids from Vernonia condensata leaves

AbstractThe aim of this study was to provide scientific knowledge to support the use of Vernonia condensata Baker, Asteraceae, beverages for their alleged hypocholesterolemic properties by testing their action as HMG-CoA reductase inhibitors and their capacity to lower dietary cholesterol permeation. Chlorogenic acid, and other caffeoylquinic acids derivatives were identified as the main components of these beverages by LC–MS/MS. No changes in the composition were notice after the in vitro gastrointestinal digestion and no toxicity against Caco-2 and HepG2 cell lines was detected. Cholesterol permeation through Caco-2 monolayers was reduced in 37% in the presence of these herbal teas, and the caffeoylquinic acids permeated the monolayers in 30–40% of their initial amount in 6h. HMG-CoA reductase activity was reduced with these beverages, showing an IC50 of 217μgml−1. It was concluded that caffeoylquinic acids, the major components, justified 98% of the enzyme inhibition measured

A step forward

Funding: Authors gratefully acknowledge Fundação para a Ciência e a Tecnologia (FCT) through projects UIDB/00645/2020, UIDB/04138/2020, PTDC/MED-QUI/31721/2017, UIDP/04138/2020 and Lusófona University, ULHT. Authors are also thankful to FCT/MCTES for the financial support to CESAM (UIDP/50017/2020, UIDB/50017/2020), through national funds.Anaplastic thyroid carcinoma (ATC) is a very rare subtype of thyroid carcinoma and one of the most lethal malignancies. Poor prognosis is mainly associated with its undifferentiated nature, inoperability, and failing to respond to the typically used therapies for thyroid cancer. Photothermal Therapy (PTT) entails using light to increase tissues’ temperature, leading to hyperthermia-mediated cell death. Tumours are more susceptible to heat as they are unable to dissipate it. By using functionalized gold nanoparticles (AuNPs) that transform light energy into heat, it is possible to target the heat to the tumour. This study aims to formulate ATC-targeted AuNPs able to convert near-infrared light into heat, for PTT of ATC. Different AuNPs were synthetized and coated. Size, morphology, and surface plasmon resonances band were determined. The optimized coated-AuNPs were then functionalized with ligands to assess ATC’s specificity. Safety, efficacy, and selectivity were assessed in vitro. The formulations were deemed safe when not irradiated (>70% cell viability) and selective for ATC. However, when irradiated, holo-transferrin-AuNPs were the most cytotoxic (22% of cell viability). The biodistribution and safety of this formulation was assessed in vivo. Overall, this novel formulation appears to be a highly promising approach to evaluate in a very near future.publishersversionpublishe

Unveiling the mechanism of action of 7α-acetoxy-6β-hydroxyroyleanone on an mrsa/visa strain: Membrane and cell wall interactions

UIDB/00100/2020 PTDC/MED-QUI/29036/2017 CEECIND/03414/2018 UIDB/04378/2020 PTDC/BIA-MIC/31645/2017 UID/DTP/04138/2019 UID/DTP/04567/2019 CBIOS/PRUID/BI1/2017 UIDB/04567/2020 UID/AMB/50017 UIDP/50017/2020 UIDB/50017/2020The number of cases of failure in the treatment of infections associated with resistant bacteria is on the rise, due to the decreasing efficacy of current antibiotics. Notably, 7α-Acetoxy-6β-hydroxyroyleanone (AHR), a diterpene isolated from different Plectranthus species, showed antibacterial activity, namely against Methicillin-resistant Staphylococcus aureus (MRSA) strains. The high antibacterial activity and low cytotoxicity render this natural compound an interesting alternative against resistant bacteria. The aim of this study is to understand the mechanism of action of AHR on MRSA, using the MRSA/Vancomycin-intermediate S. aureus (VISA) strain CIP 106760, and to study the AHR effect on lipid bilayers and on the cell wall. Although AHR interacted with lipid bilayers, it did not have a significant effect on membrane passive permeability. Alternatively, bacteria treated with this royleanone displayed cell wall disruption, without revealing cell lysis. In conclusion, the results gathered so far point to a yet undescribed mode of action that needs further investigation.publishersversionpublishe

A Step Forward in Breast Cancer Research: From a Natural-Like Experimental Model to a Preliminary Photothermal Approach

Supplementary Materials - The following are available online at https://www.mdpi.com/1422-0067/21/24/9681/s1, Figure S1: GNPs’ size distribution by intensity (%) obtained by DLS.Breast cancer is one of the most frequently diagnosed malignancies and common causes of cancer death in women. Recent studies suggest that environmental exposures to certain chemicals, such as 7,12-Dimethylbenzanthracene (DMBA), a chemical present in tobacco, may increase the risk of developing breast cancer later in life. The first-line treatments for breast cancer (surgery, chemotherapy or a combination of both) are generally invasive and frequently associated with severe side effects and high comorbidity. Consequently, novel approaches are strongly required to find more natural-like experimental models that better reflect the tumors’ etiology, physiopathology and response to treatments, as well as to find more targeted, efficient and minimally invasive treatments. This study proposes the development and an in deep biological characterization of an experimental model using DMBA-tumor-induction in Sprague-Dawley female rats. Moreover, a photothermal therapy approach using a near-infrared laser coupled with gold nanoparticles was preliminarily assessed. The gold nanoparticles were functionalized with Epidermal Growth Factor, and their physicochemical properties and in vitro effects were characterized. DMBA proved to be a very good and selective inductor of breast cancer, with 100% incidence and inducing an average of 4.7 tumors per animal. Epigenetic analysis showed that tumors classified with worst prognosis were hypomethylated. The tumor-induced rats were then subjected to a preliminary treatment using functionalized gold nanoparticles and its activation by laser (650–900 nm). The treatment outcomes presented very promising alterations in terms of tumor histology, confirming the presence of necrosis in most of the cases. Although this study revealed encouraging results as a breast cancer therapy, it is important to define tumor eligibility and specific efficiency criteria to further assess its application in breast cancer treatment on other species.The APC was funded by Faculty of Pharmacy, University of Coimbra and Coimbra Chemistry Centre, Department of Chemistry, University of Coimbra. This work was also supported by Fundação para a Ciência e Tecnologia (FCT), Portugal under the project’s references UIDB/00645/2020 and UID/DTP/04138/2019. TFG was supported by FCT, Portugal under the reference SFRH/BD/147306/2019. Thanks to FCT/MCTES for the financial support to CESAM (UIDP/50017/2020+UIDB/50017/2020), through national funds.info:eu-repo/semantics/publishedVersio

How can biomolecules improve mucoadhesion of oral insulin? A comprehensive insight using ex-vivo, in silico and in vivo models

Currently, insulin can only be administered through the subcutaneous route. Due to the flaws associated with this route, it is of interest to orally deliver this drug. However, insulin delivered orally has several barriers to overcome as it is degraded by the stomach’s low pH, enzymatic content, and poor absorption in the gastrointestinal tract. Polymers with marine source like chitosan are commonly used in nanotechnology and drug delivery due to their biocompatibility and special features. This work focuses on the preparation and characterization of mucoadhesive insulin-loaded polymeric nanoparticles. Results showed a suitable mean size for oral administration (<600 nm by dynamic laser scattering), spherical shape, encapsulation efficiency (59.8%), and high recovery yield (80.6%). Circular dichroism spectroscopy demonstrated that protein retained its secondary structure after encapsulation. Moreover, the mucoadhesive potential of the nanoparticles was assessed in silico and the results, corroborated with ex-vivo experiments, showed that using chitosan strongly increases mucoadhesion. Besides, in vitro and in vivo safety assessment of the final formulation were performed, showing no toxicity. Lastly, the insulin-loaded nanoparticles were effective in reducing diabetic rats’ glycemia. Overall, the coating of insulin-loaded nanoparticles with chitosan represents a potentially safe and promising approach to protect insulin and enhance peroral delivery.Supported in part by UID/DTP/04138/2019 from FCT, Portugal and DREAMS (ULHT). SEM analysis was funded by FCT/MCTES for the financial support to CESAM (UIDP/50017/2020+UIDB/50017/2020), through national funds. Acknowledgments: The authors are grateful to the Carla Vânia (iMedUlisboa) for her collaboration in HPLC analysis and Joana Moreira (ECTS-ULHT) for her collaboration in conducting some experiments.info:eu-repo/semantics/publishedVersio

How can biomolecules improve mucoadhesion of oral insulin? A comprehensive insight using ex-vivo, in silico, and in vivo models

Funding: Supported in part by UID/DTP/04138/2019 from FCT, Portugal and DREAMS (ULHT). SEM analysis was funded by FCT/MCTES for the financial support to CESAM (UIDP/50017/2020+UIDB/50017/2020), through national funds.Currently, insulin can only be administered through the subcutaneous route. Due to the flaws associated with this route, it is of interest to orally deliver this drug. However, insulin delivered orally has several barriers to overcome as it is degraded by the stomach’s low pH, enzymatic content, and poor absorption in the gastrointestinal tract. Polymers with marine source like chitosan are commonly used in nanotechnology and drug delivery due to their biocompatibility and special features. This work focuses on the preparation and characterization of mucoadhesive insulin-loaded polymeric nanoparticles. Results showed a suitable mean size for oral administration (<600 nm by dynamic laser scattering), spherical shape, encapsulation efficiency (59.8%), and high recovery yield (80.6%). Circular dichroism spectroscopy demonstrated that protein retained its secondary structure after encapsulation. Moreover, the mucoadhesive potential of the nanoparticles was assessed in silico and the results, corroborated with ex-vivo experiments, showed that using chitosan strongly increases mucoadhesion. Besides, in vitro and in vivo safety assessment of the final formulation were performed, showing no toxicity. Lastly, the insulin-loaded nanoparticles were effective in reducing diabetic rats’ glycemia. Overall, the coating of insulin-loaded nanoparticles with chitosan represents a potentially safe and promising approach to protect insulin and enhance peroral delivery.publishersversionpublishe

Further evidence of possible therapeutic uses of Sambucus nigra L. extracts by the assessment of the In Vitro and In Vivo anti-inflammatory properties of Its PLGA and PCL-Based Nanoformulations

Sambucus nigra L. is widely used in traditional medicine with different applications. However, confirmative studies are strongly required. This study aimed to assess the biological activities of the S. nigra flower’s extract encapsulated into two different types of nanoparticles for optimizing its properties and producing further evidence of its potential therapeutic uses. Different nanoparticles (poly(lactide-co-glycolide, PLGA) and poly-Ɛ-caprolactone (PCL), both with oleic acid, were prepared by emulsification/solvent diffusion and solvent-displacement methods, respectively. Oleic acid was used as a capping agent. After the nanoparticles’ preparation, they were characterized and the biological activities were studied in terms of collagenase, in vitro and in vivo anti-inflammatory, and in vitro cell viability. Rutin and naringenin were found to be the major phenolic compounds in the studied extract. The encapsulation efficiency was higher than 76% and revealed to have an impact on the release of the extract, mainly for the PLGA. Moreover, biochemical and histopathological analyses confirmed that the extract-loaded PLGA-based nanoparticles displayed the highest anti-inflammatory activity. In addition to supporting the previously reported evidence of potential therapeutic uses of S. nigra, these results could draw the pharmaceutical industry’s interest to the novelty of the nanoproducts.Authors also gratefully acknowledge the Régiefrutas and Fundação para a Ciência e a Tecnologia (FCT) (UIDB/04138/2020, UIDB/00100/2020, UIDP/50017/2020+UIDB/50017/2020) and Portugal 2020 to the Portuguese Mass Spectrometry Network (LISBOA-01-0145-FEDER-402-022125) for financial support. Furthermore, the authors are also grateful to the work supported by the FCT and the Portuguese National and Regional Budget, through CCMAR/Multi/04326/2019 project. Finally, Luísa Custódio was supported by the FCT Scientific Employment Stimulus (CEECIND/00425/2017).info:eu-repo/semantics/publishedVersio