Antimicrobial, Antioxidant, and Anti-Inflammatory Properties of Monofloral Honeys from Chile.

Honey is a mixture of compounds produced by bees that has been appreciated by humanity since the creation of the oldest civilizations. It has multiple uses and can be a highly nutritional and healing substance. It has been used in traditional medicine as a natural alternative for the treatment of diverse clinical conditions. This is due to its reported bioactive properties. The objective of this article is to exhibit and analyze the biological properties of different types of honey originating from Chile based on their antimicrobial, antioxidant, and anti-inflammatory activities, focusing primarily on recompiling experimental studies made on monofloral honey of plant species present in the Chilean territory. The result of this bibliographical review shows that Chilean honey possesses remarkable bioactive properties, mainly antimicrobial and antioxidant activities, with a few studies on its anti-inflammatory activity. Most of these results were attributed to monofloral honey belonging to ulmo (Eucryphia cordifolia) and quillay (Quillaja saponaria Molina) plant species. These properties are related to the presence of several bioactive components, such as phenolic components (mainly flavonoids), hydrogen peroxide (H2O2), enzymes, proteins, and carbohydrates. The biodiversity of the flora and the environmental conditions of the Chilean territory are responsible for the wide range of bioactive compounds and biological properties found in Chilean honey. Further studies must be made to uncover the medicinal potential of these native honeys.

Statins and Hemostasis: Therapeutic Potential Based on Clinical Evidence.

Hemostasis preserves blood fluidity and prevents its loss after vessel injury. The maintenance of blood fluidity requires a delicate balance between pro-coagulant and fibrinolytic status. Endothelial cells (ECs) in the inner face of blood vessels maintain hemostasis through balancing anti-thrombotic and pro-fibrinolytic activities. Dyslipidemias are linked to hemostatic alterations. Thus, it is necessary a better understanding of the underlying mechanisms linking hemostasis with dyslipidemia. Statins are drugs that decrease cholesterol levels in the blood and are the gold standard for treating hyperlipidemias. Statins can be classified into natural and synthetic molecules, approved for the treatment of hypercholesterolemia. The classical mechanism of action of statins is by competitive inhibition of a key enzyme in the synthesis pathway of cholesterol, the HMG-CoA reductase. Statins are frequently administrated by oral ingestion and its interaction with other drugs and food supplements is associated with altered bioavailability. In this review we deeply discuss the actions of statins beyond the control of dyslipidemias, focusing on the actions in thrombotic modulation, vascular and cardiovascular-related diseases, metabolic diseases including metabolic syndrome, diabetes, hyperlipidemia, and hypertension, and chronic diseases such as cancer, chronic obstructive pulmonary disease, and chronic kidney disease. Furthermore, we were prompted to delved deeper in the molecular mechanisms by means statins regulate coagulation acting on liver, platelets, and endothelium. Clinical evidence show that statins are effective regulators of dyslipidemia with a high impact in hemostasis regulation and its deleterious consequences. However, studies are required to elucidate its underlying molecular mechanism and improving their therapeutical actions.

α1-Adrenergic Stimulation Increases Platelet Adhesion to Endothelial Cells Mediated by TRPC6.

Stimulation of a1-adrenergic nervous system is increased during systemic inflammation and other pathological conditions with the consequent adrenergic receptors (ARs) activation. It has been reported that a1-stimulation contributes to coagulation since a1-AR blockers inhibit coagulation and its organic consequences. Also, coagulation induced by a1-AR stimulation can be greatly decreased using a1-AR blockers. In health, endothelial cells (ECs) perform anticoagulant actions at cellular and molecular level. However, during inflammation, ECs turn dysfunctional promoting a procoagulant state. Endothelium-dependent coagulation progresses at cellular and molecular levels, promoting endothelial acquisition of procoagulant properties to potentiate coagulation by means of prothrombotic and antifibrinolytic proteins expression increase in ECs releasing them to circulation, the thrombus formation is strengthened. Calcium signaling is a main feature of coagulation. Inhibition of ion channels involved in Ca2+ entry severely decreases coagulation. The transient receptor potential canonical 6 (TRPC6) is a non-selective Ca2+-permeable ion channel. TRPC6 activity is induced by diacylglycerol, suggesting that is regulated by a1-ARs. Furthermore, a1-ARs stimulation elicits a TRPC-like current in rat mesenteric artery smooth muscle and mesangial cells. However, whether TRPC6 could promote an ECs-mediated platelet adhesion induced by a1-adrenergic stimulation is currently not known. Therefore, the aim of this study was to examine if the TRPC6 calcium channel mediates platelet adhesion induced by a1-adrenergic stimulation. Our results suggest that platelet adhesion to ECs is enhanced by the a1-adrenergic stimulation evoked by phenylephrine mediated by TRPC6 activity. We conclude that TRPC6 is a molecular determinant in platelet adhesion to ECs with implications in systemic inflammatory diseases treatment.

Anti-Inflammatory Chilean Endemic Plants.

Medicinal plants have been used since prehistoric times and continue to treat several diseases as a fundamental part of the healing process. Inflammation is a condition characterized by redness, pain, and swelling. This process is a hard response by living tissue to any injury. Furthermore, inflammation is produced by various diseases such as rheumatic and immune-mediated conditions, cancer, cardiovascular diseases, obesity, and diabetes. Hence, anti-inflammatory-based treatments could emerge as a novel and exciting approach to treating these diseases. Medicinal plants and their secondary metabolites are known for their anti-inflammatory properties, and this review introduces various native Chilean plants whose anti-inflammatory effects have been evaluated in experimental studies. Fragaria chiloensis, Ugni molinae, Buddleja globosa, Aristotelia chilensis, Berberis microphylla, and Quillaja saponaria are some native species analyzed in this review. Since inflammation treatment is not a one-dimensional solution, this review seeks a multidimensional therapeutic approach to inflammation with plant extracts based on scientific and ancestral knowledge.

Sepsis-Induced Coagulopathy Phenotype Induced by Oxidized High-Density Lipoprotein Associated with Increased Mortality in Septic-Shock Patients.

Sepsis syndrome is a highly lethal uncontrolled response to an infection, which is characterized by sepsis-induced coagulopathy (SIC). High-density lipoprotein (HDL) exhibits antithrombotic activity, regulating coagulation in vascular endothelial cells. Sepsis induces the release of several proinflammatory molecules, including reactive oxygen species, which lead to an increase in oxidative stress in blood vessels. Thus, circulating lipoproteins, such as HDL, are oxidized to oxHDL, which promotes hemostatic dysfunction, acquiring prothrombotic properties linked to the severity of organ failure in septic-shock patients (SSP). However, a rigorous and comprehensive investigation demonstrating that oxHDL is associated with a coagulopathy-associated deleterious outcome of SSP, has not been reported. Thus, we investigated the participation of plasma oxHDL in coagulopathy-associated sepsis pathogenesis and elucidated the underlying molecular mechanism. A prospective study was conducted on 42 patients admitted to intensive care units, (26 SSP and 16 non-SSP) and 39 healthy volunteers. We found that an increased plasma oxHDL level in SSP was associated with a prothrombotic phenotype, increased mortality and elevated risk of death, which predicts mortality in SSP. The underlying mechanism indicates that oxHDL triggers an endothelial protein expression reprogramming of coagulation factors and procoagulant adhesion proteins, to produce a prothrombotic environment, mainly mediated by the endothelial LOX-1 receptor. Our study demonstrates that an increased plasma oxHDL level is associated with coagulopathy in SSP through a mechanism involving the endothelial LOX-1 receptor and endothelial protein expression regulation. Therefore, the plasma oxHDL level plays a role in the molecular mechanism associated with increased mortality in SSP.

Procoagulant phenotype induced by oxidized high-density lipoprotein associates with acute kidney injury and death.

BACKGROUND: Oxidative stress derived from severe systemic inflammation promotes conversion from high-density lipoprotein HDL to oxidized HDL (oxHDL), which interacts with vascular endothelial cells (ECs). OxHDL acquires procoagulant features playing a role in modulating coagulation, which has been linked with organ failure in ICU patients. However, whether oxHDL elicits a ECs-mediated procoagulant phenotype generating organ failure and death, and the underlying molecular mechanism is not known. Therefore, we studied whether oxHDL-treated rats and high-oxHDL ICU patients exhibit a procoagulant phenotype and its association with kidney injury and mortality and the endothelial underlying molecular mechanism. METHODS: Human ECs, oxHDL-treated rats and ICU patients were subjected to several cellular and molecular studies, coagulation analyses, kidney injury assessment and mortality determination. RESULTS: OxHDL-treated ECs showed a procoagulant protein expression reprograming characterized by increased E-/P-selectin and vWF mRNA expression through specific signaling pathways. OxHDL-treated rats exhibited a procoagulant phenotype and modified E-/P-selectin, vWF, TF and t-PA mRNA expression correlating with plasma TF, t-PA and D-dimer. Also, showed increased death events and the relative risk of death, and increased creatinine, urea, BUN/creatinine ratio, KIM-1, NGAL, ß2M, and decreased eGFR, all concordant with kidney injury, correlated with plasma TF, t-PA and D-dimer. ICU patients showed correlation between plasma oxHDL and increased creatinine, cystatin, BUN, BUN/creatinine ratio, KIM-1, NGAL, ß2M, and decreased GFR. Notably, ICU high-oxHDL patients showed decreased survival. Interestingly, altered coagulation factors TF, t-PA and D-dimer correlated with both increased oxHDL levels and kidney injury markers, indicating a connection between these factors. CONCLUSION: Increased circulating oxHDL generates an endothelial-dependent procoagulant phenotype that associates with acute kidney injury and increased risk of death.

Biochemical characterization of Peumus boldus fruits: Insights of its antioxidant properties through a theoretical approach.

Peumus boldus is an endemic tree species from Chile whose leaves have been the focus of study for decades given that their infusions are reported to relieve rheumatic symptoms, headache, dyspepsia, urinary tract inflammation, and symptoms of other illnesses. These health properties have been studied mainly using leaves and bark, then it is relevant to know more about these properties in different parts of the plant. Considering the importance of P. boldus fruits in the diet of some rural populations, we analyzed their properties to explore its impact on the Chilean population health. Liquid chromatography and mass spectrometry analysis confirmed the presence of alkaloids such as boldine, although aporphine N-methyl-laurotetanine was the most abundant. In addition, flavonoids catechin, chrysin and quercetin were also found in the extract. Cytotoxicity and anti-inflammatory activities of the fruit extract were invitro tested by using a murine macrophage cell model, observing that a diluted fraction of the extract was not cytotoxic, but showed anti-inflammatory activity, which is likely attributed to antioxidants activities. By means of quantum chemical calculations, we calculated the redox potential of the respective alkaloids and flavonoids found in the extract. Results suggest a synergistic effect between alkaloids and flavonoids, where boldine and N-methyl-laurotetanine showed similar antioxidant properties. Finally, we present a description of the oxidation mechanisms for both groups of molecules which will sustain P. boldus fruit biological properties, in order to give this kind of fruits scientific value focusing on human health.

Development of a PHBV nanoparticle as a peptide vehicle for NOD1 activation.

NOD1 is an intracellular receptor that, when activated, induces gene expression of pro-inflammatory factors promoting macrophages and neutrophils recruitment at the infection site. However, iE-DAP, the dipeptide agonist that promotes this receptor's activation, cannot permeate cell membranes. To develop a nanocarrier capable of achieving a high and prolonged activation over time, iE-DAP was encapsulated in nanoparticles (NPs) made of poly (3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV). The physicochemical properties, colloidal stability, encapsulation efficiency, and cellular uptake of iE-DAP-loaded PHVB NPs were analyzed. Results evidenced that physicochemical properties of iE-DAP-loaded NPs remained stable over time, and NPs were efficiently internalized into cells, a process that depends on time and concentration. Moreover, our results showed that NPs elicited a controlled cargo release in vitro, and the encapsulated agonist response was higher than its free form, suggesting the possibility of activating intracellular receptors triggering an immune response through the release of NOD1 agonist.

Rhenium (I) Complexes as Probes for Prokaryotic and Fungal Cells by Fluorescence Microscopy: Do Ligands Matter?

Re(I) complexes have exposed highly suitable properties for cellular imaging (especially for fluorescent microscopy) such as low cytotoxicity, good cellular uptake, and differential staining. These features can be modulated or tuned by modifying the ligands surrounding the metal core. However, most of Re(I)-based complexes have been tested for non-walled cells, such as epithelial cells. In this context, it has been proposed that Re(I) complexes are inefficient to stain walled cells (i.e., cells protected by a rigid cell wall, such as bacteria and fungi), presumably due to this physical barrier hampering cellular uptake. More recently, a series of studies have been published showing that a suitable combination of ligands is useful for obtaining Re(I)-based complexes able to stain walled cells. This review summarizes the main characteristics of different fluorophores used in bioimage, remarking the advantages of d6-based complexes, and focusing on Re(I) complexes. In addition, we explored different structural features of these complexes that allow for obtaining fluorophores especially designed for walled cells (bacteria and fungi), with especial emphasis on the ligand choice. Since many pathogens correspond to bacteria and fungi (yeasts and molds), and considering that these organisms have been increasingly used in several biotechnological applications, development of new tools for their study, such as the design of new fluorophores, is fundamental and attractive.

New Properties of a Bioinspired Pyridine Benzimidazole Compound as a Novel Differential Staining Agent for Endoplasmic Reticulum and Golgi Apparatus in Fluorescence Live Cell Imaging.

In this study, we explored new properties of the bioinspired pyridine benzimidazole compound B2 (2,4-di-tert-butyl-6-(3H-imidazo[4,5-c]pyridine-2-yl)phenol) regarding its potential use as a differential biomarker. For that, we performed 1D 1HNMR (TOCSY), UV-Vis absorption spectra in different organic solvents, voltammetry profile (including a scan-rate study), and TD-DFT calculations that including NBO analyses, to provide valuable information about B2 structure and luminescence. In our study, we found that the B2 structure is highly stable, where the presence of an intramolecular hydrogen bond (IHB) seems to have a crucial role in the stability of luminescence, and its emission can be assigned as fluorescence. In fact, we found that the relatively large Stokes Shift observed for B2 (around 175 nm) may be attributed to the stability of the B2 geometry and the strength of its IHB. On the other hand, we determined that B2 is biocompatible by cytotoxicity experiments in HeLa cells, an epithelial cell line. Furthermore, in cellular assays we found that B2 could be internalized by passive diffusion in absence of artificial permeabilization at short incubation times (15 min to 30 min). Fluorescence microscopy studies confirmed that B2 accumulates in the endoplasmic reticulum (ER) and Golgi apparatus, two organelles involved in the secretory pathway. Finally, we determined that B2 exhibited no noticeable blinking or bleaching after 1 h of continuous exposure. Thus, B2 provides a biocompatible, rapid, simple, and efficient way to fluorescently label particular organelles, producing similar results to that obtained with other well-established but more complex methods.

Intracellular trafficking and cellular uptake mechanism of PHBV nanoparticles for targeted delivery in epithelial cell lines.

BACKGROUND: Nanotechnology is a science that involves imaging, measurement, modeling and a manipulation of matter at the nanometric scale. One application of this technology is drug delivery systems based on nanoparticles obtained from natural or synthetic sources. An example of these systems is synthetized from poly(3-hydroxybutyrate-co-3-hydroxyvalerate), which is a biodegradable, biocompatible and a low production cost polymer. The aim of this work was to investigate the uptake mechanism of PHBV nanoparticles in two different epithelial cell lines (HeLa and SKOV-3). RESULTS: As a first step, we characterized size, shape and surface charge of nanoparticles using dynamic light scattering and transmission electron microscopy. Intracellular incorporation was evaluated through flow cytometry and fluorescence microscopy using intracellular markers. We concluded that cellular uptake mechanism is carried out in a time, concentration and energy dependent way. Our results showed that nanoparticle uptake displays a cell-specific pattern, since we have observed different colocalization in two different cell lines. In HeLa (Cervical cancer cells) this process may occur via classical endocytosis pathway and some internalization via caveolin-dependent was also observed, whereas in SKOV-3 (Ovarian cancer cells) these patterns were not observed. Rearrangement of actin filaments showed differential nanoparticle internalization patterns for HeLa and SKOV-3. Additionally, final fate of nanoparticles was also determined, showing that in both cell lines, nanoparticles ended up in lysosomes but at different times, where they are finally degraded, thereby releasing their contents. CONCLUSIONS: Our results, provide novel insight about PHBV nanoparticles internalization suggesting that for develop a proper drug delivery system is critical understand the uptake mechanism.