Differential lipid accumulation on HepG2 cells triggered by palmitic and linoleic fatty acids exposure

Lipid metabolism pathways such as β-oxidation, lipolysis and, lipogenesis, are mainly associated with normal liver function. However, steatosis is a growing pathology caused by the accumulation of lipids in hepatic cells due to increased lipogenesis, dysregulated lipid metabolism, and/or reduced lipolysis. Accordingly, this investigation hypothesizes a selective in vitro accumulation of palmitic and linoleic fatty acids on hepatocytes. After assessing the metabolic inhibition, apoptotic effect, and reactive oxygen species (ROS) generation by linoleic (LA) and palmitic (PA) fatty acids, HepG2 cells were exposed to different ratios of LA and PA to study the lipid accumulation using the lipophilic dye Oil Red O. Lipidomic studies were also carried out after lipid isolation. Results revealed that LA was highly accumulated and induced ROS production when compared to PA. Lipid profile modifications were observed after LA:PA 1:1 (v/v) exposure, which led to a four-fold increase in triglycerides (TGs) (mainly in linoleic acid-containing species), as well as a increase in cholesterol and polyunsaturated fatty acids (PUFA) content when compared to the control cells. The present work highlights the importance of balancing both PA and LA fatty acids concentrations in HepG2 cells to maintain normal levels of free fatty acids (FFAs), cholesterol, and TGs and to minimize some of the observed in vitro effects (i.e., apoptosis, ROS generation and lipid accumulation) caused by these fatty acids.info:eu-repo/semantics/publishedVersio

Cannabidiol and cannabigerol exert antimicrobial activity without compromising skin microbiota

Cannabidiol (CBD) and cannabigerol (CBG) are two pharmacologically active phytocannabinoids of Cannabis sativa L. Their antimicrobial activity needs further elucidation, particularly for CBG, as reports on this cannabinoid are scarce. We investigated CBD and CBG’s antimicrobial potential, including their ability to inhibit the formation and cause the removal of biofilms. Our results demonstrate that both molecules present activity against planktonic bacteria and biofilms, with both cannabinoids removing mature biofilms at concentrations below the determined minimum inhibitory concentrations. We report for the first time minimum inhibitory and lethal concentrations for Pseudomonas aeruginosa and Escherichia coli (ranging from 400 to 3180 µM), as well as the ability of cannabinoids to inhibit Staphylococci adhesion to keratinocytes, with CBG demonstrating higher activity than CBD. The value of these molecules as preservative ingredients for cosmetics was also assayed, with CBG meeting the USP 51 challenge test criteria for antimicrobial effectiveness. Further, the exact formulation showed no negative impact on skin microbiota. Our results suggest that phytocannabinoids can be promising topical antimicrobial agents when searching for novel therapeutic candidates for different skin conditions. Additional research is needed to clarify phytocannabinoids’ mechanisms of action, aiming to develop practical applications in dermatological use.info:eu-repo/semantics/publishedVersio

Particle size effect of integral carob flour on bioaccessibility of bioactive compounds during simulated gastrointestinal digestion

Carob fruit is native to the Mediterranean region and produced mainly in Portugal, Italy, Morocco and Turkey. The production of the carob fruit in Portugal is highly extensive and sustainable. Currently, carob flour (CF) production is mainly achieved after pulp separation, despite it having been demonstrated that the seeds improve the extraction efficiency of bioactive compounds such as polyphenols, promoting human health. This study aimed to produce an integral CF through an innovative process and assess its physicochemical and bioactive properties at different particle sizes throughout simulated gastrointestinal tract (GIT) digestion. The sugar content profile obtained throughout GIT digestion indicated that sucrose, the sugar present at the highest concentration in undigested CF, was digested and broken down into simple sugars, namely glucose and fructose. The total phenolic content (TPC) and antioxidant activity obtained for the ≤100 µm fraction were in accordance and gastric digestion promoted an increase in the TPC value compared to the undigested sample. The >100 µm fractions displayed a distinct profile from the ≤100 µm fraction. This study showed that the particle size affects the sugar, antioxidant and total phenolic content of CFs and also their gastrointestinal tract digestion. The ≤100 µm fraction demonstrated the most suitable profile as a functional food ingredient.info:eu-repo/semantics/publishedVersio

Development of an inhalational therapeutical system based on bacteriophages to treat pharyngo-tonsillitis : a nanoencapsulation approach

Inflammatory diseases that occur in the pharynx and involving both the adenoids and tonsils are important not only for being very frequent, but also because they often require minor surgery for their resolution. These structures have immunological functions leading to production of antibodies, and work in the local immunity of the pharynx and protection of the entire body. The most common etiologic agent of sore throats is Streptococcus pyogenes, an important pathogen of the beta-hemolytic group A which causes streptococcal pharyngitis. The emergence of antibiotic-resistant bacterial strains and the poor penetration of chemical antibiotics in bacterial biofilms raise the need for safe and effective options of antimicrobial treatment. The application of bacteriophages (or cocktails therefrom) has been proposed as an alternative (or complement) to conventional chemical antibiotics, allowing the release of natural predators of bacteria directly on these biofilms. The major advantage of bacteriophage-based antibiotherapy relative to its conventional chemical counterpart is that bacteriophages replicate at the site of infection, being available in abundance where they are needed the most. When compared with chemical antibiotics, bacteriophages have other important advantages: (i) strong tissue permeability, (ii) bacteriophage concentration remains high at the focus of infection, continuously increasing with bacterial (host) presence, (iii) elimination of the focus of infection occurs only after eradication of the host bacterium, (iv) bacteriophages are fully compatible with antibiotics and may act synergistically, (v) they are specific against the target bacteria, (vi) have a superior ability to penetrate bacterial biofilms, inducing production of enzymes that hydrolyze the biofilm polymeric matrix, (vii) although bacteria can develop resistance to bacteriophages, isolation of new lytic bacteriophages is much simpler and cheaper than developing a new chemical antibiotic. In this research effort, development of a biotechnological process for the inhalational administration of a bacteriophage cocktail (endotoxin free) was pursued, using strategies of nanoencapsulation within lipid nanovesicles (as forms of protection for the bacteriophage against the immune system) to treat infectious pathologies such as pharyngo-tonsillitis caused by Streptococcus pyogenes. This method of targeting may have a high potential for the treatment of bacterial infections of the respiratory tract, since inhalation therapy is considered to be favorable to certain respiratory infections because the aerosol is delivered directly at the site of infection, accelerating the action of bacterial predators. Additionally, a smaller amount of bioactive substance is needed, thus preventing or reducing possible side effects. As a proof of concept for the nanoencapsulation strategy, and since there is not yet available a strictly lytic bacteriophage cocktail for Streptococcus pyogenes, a well-defined and characterized bacteriophage was utilized, viz. bacteriophage T4. Water-in-oil-in-water (W/O/W) multiple emulsions are nanosystems in which dispersions of small water droplets within larger oil droplets are themselves dispersed in a continuous aqueous phase. Due to their compartimentalized internal structure, multiple emulsions present important advantages over simple O/W emulsions for encapsulation of biomolecules, such as the ability to carry both polar and non-polar molecules, and a better control over releasing of therapeutic molecules. T4 bacteriophage was entrapped within W/O/W multiple nanoemulsions, aiming at mimicking the multifunctional design of biology, optimized with several lipid matrices, poloxamers and stabilizing layer compositions. Physicochemical characterization of the optimized bacteriophageencasing nanovesicle formulations encompassed determination of particle size, size distribution and particle charge, via Zeta potential analysis, surface morphology via CRYO-SEM, and thermal analysis via DSC

Morphological changes in Bombyx Mori silk gland and gut, in association with the feeding of iron oxide nanoparticles

The Bombyx mori silkworm is very important econom- ically and is a model for several studies. The larval life of silkworms is divided into five instars. The 5th instar is the longest, where the larvae show maximum food consumption and growth. The silk is produced in a paired gland composed of glandular epithelium and a lumen. Food digestion occurs in the midgut, represented by a folded columnar monolayered epithelium, which includes goblet cells and stem cells in a basal lamina. We aimed to investigate the influence of concentrations of iron oxide nanoparticles (IONPs) in food on the silk gland and midgut morphology

Phytosterols and novel triterpenes recovered from industrial fermentation coproducts exert in vitro anti‐inflammatory activity in macrophages

The unstoppable growth of human population that occurs in parallel with all manufacturing activities leads to a relentless increase in the demand for resources, cultivation land, and energy. In response, currently, there is significant interest in developing strategies to optimize any available resources and their biowaste. While solutions initially focused on recovering biomolecules with applications in food, energy, or materials, the feasibility of synthetic biology in this field has been demonstrated in recent years. For instance, it is possible to genetically modify Saccharomyces cerevisiae to produce terpenes for commercial applications (i.e., against malaria or as biodiesel). But the production process, similar to any industrial activity, generates biowastes containing promising biomolecules (from fermentation) that if recovered may have applications in different areas. To test this hypothesis, in the present study, the lipid composition of by‐products from the industrial production of β‐farnesene by genetically modified Saccharomyces cerevisiae are studied to identify potentially bioactive compounds, their recovery, and finally, their stability and in vitro bioactivity. The assayed biowaste showed the presence of triterpenes, phytosterols, and 1‐ octacosanol which were recovered through molecular distillation into a single fraction. During the assayed stability test, compositional modifications were observed, mainly for the phytosterols and 1‐octacosanol, probably due to oxidative reactions. However, such changes did not affect the in vitro bioactivity in macrophages, where it was found that the obtained fraction decreased the production of TNF‐α and IL‐6 in lipopolysaccharide (LPS)‐induced inflammation.info:eu-repo/semantics/publishedVersio

Novel hydroxyapatite/chitosan bilayered scaffold for osteochondral tissue-engineering applications : scaffold design and its performance when seeded with goat bone marrow stromal cells

Recent studies suggest that bone marrow stromal cells are a potential source of osteoblasts and chondrocytes and can be used to regenerate damaged tissues using a tissue-engineering (TE) approach. However, these strategies require the use of an appropriate scaffold architecture that can support the formation de novo of either bone and cartilage tissue, or both, as in the case of osteochondral defects. The later has been attracting a great deal of attention since it is considered a difficult goal to achieve. This work consisted on developing novel hydroxyapatite/chitosan (HA/CS) bilayered scaffold by combining a sintering and a freeze-drying technique, and aims to show the potential of such type of scaffolds for being used in TE of osteochondral defects. The developed HA/CS bilayered scaffolds were characterized by Fourier transform infra-red spectroscopy, X-ray diffraction analysis, micro-computed tomography, and scanning electron microscopy (SEM). Additionally, the mechanical properties of HA/CS bilayered scaffolds were assessed under compression. In vitro tests were also carried out, in order to study the water-uptake and weight loss profile of the HA/CS bilayered scaffolds. This was done by means of soaking the scaffolds into a phosphate buffered saline for 1 up to 30 days. The intrinsic cytotoxicity of the HA scaffolds and HA/CS bilayered scaffolds extract fluids was investigated by carrying out a cellular viability assay (MTS test) using Mouse fibroblastic-like cells. Results have shown that materials do not exert any cytotoxic effect. Complementarily, in vitro (phase I) cell culture studies were carried out to evaluate the capacity of HA and CS layers to separately, support the growth and differentiation of goat marrow stromal cells (GBMCs) into osteoblasts and chondrocytes, respectively. Cell adhesion and morphology were analysed by SEM while the cell viability and proliferation were assessed by MTS test and DNA quantification. The chondrogenic differentiation of GBMCs was evaluated measuring the glucosaminoglycans synthesis. Data showed that GBMCs were able to adhere, proliferate and osteogenic differentiation was evaluated by alkaline phosphatase activity and immunocytochemistry assays after 14 days in osteogenic medium and into chondrocytes after 21 days in culture with chondrogenic medium. The obtained results concerning the physicochemical and biological properties of the developed HA/CS bilayered scaffolds, show that these constructs exhibit great potential for their use in TE strategies leading to the formation of adequate tissue substitutes for the regeneration of osteochondral defects