Desarrollo de un nanosistema para la entrega selectiva de microRNAs hacia células de cáncer ovárico

El cáncer ovárico (CO) es la patología ginecológica más letal en las mujeres. Está ligada a un bajo pronóstico de vida dado el alto porcentaje de diagnósticos tardíos. Se caracteriza por presentar tumores voluminosos, altamente irrigados y con importante diseminación peritoneal. La quimiorresistencia y recidivas post cirugía son frecuentes. Uno de los acercamientos terapéuticos más interesantes es la restauración de microRNAs (miRs) downregulados durante la carcinogénesis. Los miRs juegan un importante rol en la multiregulación de procesos celulares como proliferación, diferenciación, supervivencia, apoptosis, ciclo celular, angiogénesis y metástasis, entre otros. En este trabajo se diseñaron y sintetizaron nanopartículas de oro transportadoras del miR-145. Para esto, fueron funcionalizadas con ácido fólico o el péptido FSH33 y cargadas con el miR-145 como agente activo. El nanosistema construido ingresó selectivamente hacia las células e inhibió significativamente la viabilidad, migración y el potencial proliferativo y clonogénico. Además, disminuyó la liberación de VEGF y redujo el tamaño de esferoides tumorales mediante daño en la membrana celular, disminución de la viabilidad y activación de la apoptosis celular. Estos resultados entregan importantes avances para el desarrollo de terapias basadas en miRs mediante la utilización de nanotecnologías y sienta las bases para su evaluación en modelos in vivoOvarian cancer (OC) is the most lethal gynecological pathology in women and is linked to a low life prognosis due to the high percentage of late diagnosis. It is characterized by large tumors, highly irrigated, with significant dissemination in the peritoneal area. After debulking surgery, a high rate of chemoresistance and relapses are frequent. One of the most interesting therapeutic approaches represents the restoration of microRNAs (miRs) which are downregulated during the carcinogenesis. miRs play an important muti-regulatory role in cellular processes such as proliferation, differentiation, survival, apoptosis, cell cycle, angiogenesis, and metastasis, among others. In this work, gold nanoparticles carrying miR-145 were designed and synthesized. First, the nanoparticles were functionalized with folic acid or the FSH33 peptide, and then loaded with miR-145, a reported agent active against OC cell lines. The constructed nanosystem selectively entered these cells and significantly inhibited cell viability, migration, and proliferative and clonogenic potential. Moreover, it diminished the release of VEGF and reduced the size of OC spheroids through the damage of cell membranes, decrease of cell viability, and activation of apoptosis. These results provide important advances in the development of miR-based therapies using nanotechnologies as selective vectors and gives the approaches for the in vivo evaluatio

Minimal information for studies of extracellular vesicles (MISEV2023): From basic to advanced approaches

Extracellular vesicles (EVs), through their complex cargo, can reflect the state of their cell of origin and change the functions and phenotypes of other cells. These features indicate strong biomarker and therapeutic potential and have generated broad interest, as evidenced by the steady year-on-year increase in the numbers of scientific publications about EVs. Important advances have been made in EV metrology and in understanding and applying EV biology. However, hurdles remain to realising the potential of EVs in domains ranging from basic biology to clinical applications due to challenges in EV nomenclature, separation from non-vesicular extracellular particles, characterisation and functional studies. To address the challenges and opportunities in this rapidly evolving field, the International Society for Extracellular Vesicles (ISEV) updates its 'Minimal Information for Studies of Extracellular Vesicles', which was first published in 2014 and then in 2018 as MISEV2014 and MISEV2018, respectively. The goal of the current document, MISEV2023, is to provide researchers with an updated snapshot of available approaches and their advantages and limitations for production, separation and characterisation of EVs from multiple sources, including cell culture, body fluids and solid tissues. In addition to presenting the latest state of the art in basic principles of EV research, this document also covers advanced techniques and approaches that are currently expanding the boundaries of the field. MISEV2023 also includes new sections on EV release and uptake and a brief discussion of in vivo approaches to study EVs. Compiling feedback from ISEV expert task forces and more than 1000 researchers, this document conveys the current state of EV research to facilitate robust scientific discoveries and move the field forward even more rapidly

Development of a Nanostructured Lipid Carrier (NLC) by a Low-Energy Method, Comparison of Release Kinetics and Molecular Dynamics Simulation

Lipid nanocarriers have a great potential for improving the physicochemical characteristics and behavior of poorly water-soluble drugs, such as aqueous dispersibility and oral bioavailability. This investigation presents a novel nanostructured lipid carrier (NLC) based on a mixture of solid lipid glycerides, fatty acid esters of PEG 1500 (Gelucire® 44/14), and an oil mix composed of capric and caprylic triglycerides (Miglyol® 812). These NLCs were developed by a simple low-energy method based on melt emulsification to yield highly encapsulating and narrowly distributed nanoparticles (~100 nm, PdI = 0.1, and zeta potential = ~−10 mV). Rhodamine 123 was selected as a poorly water-soluble drug model and owing to its spectroscopic properties. The novel NLCs were characterized by dynamic light scattering (DLS), zeta potential, nanoparticle tracking analysis (NTA), transmission electron microscopy (TEM), differential scanning calorimetry (DSC), and colloidal stability. The drug release was determined through a dialysis bag and vertical Franzs’ cells to provide insights about the methods’ suitability, revealing similar performance regardless of their different fluid dynamics. Rhodamine 123 followed a characteristic biphasic release profile owing to the swelling of the hydrophilic polymer coating and diffusion process from the lipid core as revealed by the Korsmeyers–Peppas kinetic modeling. Moreover, to elucidate the formation and incorporation of Rhodamine 123 into the NLC core, several molecular dynamics simulations were conducted. The temperature was shown to be an important condition to improve the formation of the nanoparticles. In addition, the liquid lipid incorporation to the formulation forms nanoparticles with imperfect centers, in contrast to nanoparticles without it. Moreover, Miglyol® 812 improves hydrophobic molecule solubility. These results suggest the potential of novel NLC as a drug delivery system for poorly water-soluble drugs.</jats:p

Development of a Nanostructured Lipid Carrier (NLC) by a Low-Energy Method, Comparison of Release Kinetics and Molecular Dynamics Simulation

Lipid nanocarriers have a great potential for improving the physicochemical characteristics and behavior of poorly water-soluble drugs, such as aqueous dispersibility and oral bioavailability. This investigation presents a novel nanostructured lipid carrier (NLC) based on a mixture of solid lipid glycerides, fatty acid esters of PEG 1500 (Gelucire® 44/14), and an oil mix composed of capric and caprylic triglycerides (Miglyol® 812). These NLCs were developed by a simple low-energy method based on melt emulsification to yield highly encapsulating and narrowly distributed nanoparticles (~100 nm, PdI = 0.1, and zeta potential = ~−10 mV). Rhodamine 123 was selected as a poorly water-soluble drug model and owing to its spectroscopic properties. The novel NLCs were characterized by dynamic light scattering (DLS), zeta potential, nanoparticle tracking analysis (NTA), transmission electron microscopy (TEM), differential scanning calorimetry (DSC), and colloidal stability. The drug release was determined through a dialysis bag and vertical Franzs’ cells to provide insights about the methods’ suitability, revealing similar performance regardless of their different fluid dynamics. Rhodamine 123 followed a characteristic biphasic release profile owing to the swelling of the hydrophilic polymer coating and diffusion process from the lipid core as revealed by the Korsmeyers–Peppas kinetic modeling. Moreover, to elucidate the formation and incorporation of Rhodamine 123 into the NLC core, several molecular dynamics simulations were conducted. The temperature was shown to be an important condition to improve the formation of the nanoparticles. In addition, the liquid lipid incorporation to the formulation forms nanoparticles with imperfect centers, in contrast to nanoparticles without it. Moreover, Miglyol® 812 improves hydrophobic molecule solubility. These results suggest the potential of novel NLC as a drug delivery system for poorly water-soluble drugs

Intranasal delivery of mesenchymal stem cell-derived exosomes reduces oxidative stress and markedly inhibits ethanol consumption and post-deprivation relapse drinking.

Chronic ethanol consumption leads to brain oxidative stress and neuroinflammation, conditions known to potentiate and perpetuate each other. Several studies have shown that neuroinflammation results in increases in chronic ethanol consumption. Recent reports showed that the intra-cerebroventricular administration of mesenchymal stem cells to rats consuming alcohol chronically markedly inhibited oxidative-stress, abolished neuroinflammation and greatly reduced chronic alcohol intake and post deprivation relapselike alcohol intake. However, the intra-cerebroventricular administration of living cells is not suitable as a treatment of a chronic condition. The present study aimed at inhibiting ethanol intake by the noninvasive intranasal administration of human mesenchymal stem cell products: exosomes; microvesicles (40 to 150 nm) with marked antioxidant activity extruded from mesenchymal stem cells. The exosome membrane can fuse with the plasma membrane of cells in different tissues, thus delivering their content intracellularly. The study showed that the weekly intranasal administration of mesenchymal stem cell-derived exosomes to rats consuming alcohol chronically (i) inhibited their ethanol intake by 80% and blunted the relapselike “binge” drinking that follows an alcohol deprivation period and ethanol re-access. (ii) Intranasally administered exosomes were found in the brain within 24-hours; (iii) fully reversed both alcohol-induced hippocampal oxidative-stress, evidenced by a lower ratio of oxidized to reduced glutathione, and neuroinflammation, shown by a reduced astrocyte activation and microglial density and (iv) increased glutamate transporter GLT1 expression in nucleus accumbens, counteracting the inhibition of glutamate transporter activity, reportedly depressed under oxidative-stress conditions. Possible translational implications are envisagedVersión enviad

Peptide Targeted Gold Nanoplatform Carrying miR-145 Induces Antitumoral Effects in Ovarian Cancer Cells

One of the recent attractive therapeutic approaches for cancer treatment is restoring downregulated microRNAs. They play an essential muti-regulatory role in cellular processes such as proliferation, differentiation, survival, apoptosis, cell cycle, angiogenesis, and metastasis, among others. In this study, a gold nanoplatform (GNPF) carrying miR-145, a downregulated microRNA in many cancer types, including epithelial ovarian cancer, was designed and synthesized. For targeting purposes, the GNPF was functionalized with the FSH33 peptide, which provided selectivity for ovarian cancer, and loaded with the miR-145 to obtain the nanosystem GNPF-miR-145. The GNPF-mir-145 was selectively incorporated in A2780 and SKOV3 cells and significantly inhibited cell viability and migration and exhibited proliferative and anchor-independent growth capacities. Moreover, it diminished VEGF release and reduced the spheroid size of ovarian cancer through the damage of cell membranes, thus decreasing cell viability and possibly activating apoptosis. These results provide important advances in developing miR-based therapies using nanoparticles as selective vectors and provide approaches for in vivo evaluation

Extracellular vesicles through the blood–brain barrier: a review

Extracellular vesicles (EVs) are particles naturally released from cells that are delimited by a lipid bilayer and are unable to replicate. How the EVs cross the Blood–Brain barrier (BBB) in a bidirectional manner between the bloodstream and brain parenchyma remains poorly understood. Most in vitro models that have evaluated this event have relied on monolayer transwell or microfluidic organ-on-a-chip techniques that do not account for the combined effect of all cellular layers that constitute the BBB at different sites of the Central Nervous System. There has not been direct transcytosis visualization through the BBB in mammals in vivo, and evidence comes from in vivo experiments in zebrafish. Literature is scarce on this topic, and techniques describing the mechanisms of EVs motion through the BBB are inconsistent. This review will focus on in vitro and in vivo methodologies used to evaluate EVs transcytosis, how EVs overcome this fundamental structure, and discuss potential methodological approaches for future analyses to clarify these issues. Understanding how EVs cross the BBB will be essential for their future use as vehicles in pharmacology and therapeutics

Stability of antibacterial silver carboxylate complexes against staphylococcus epidermidis and their cytotoxic effects

© 2018 by the authors. The antibacterial effects against Staphylococcus epidermidis of five silver carboxylate complexes with anti-inflammatory ligands were studied in order to analyze and compare them in terms of stability (in solution and after exposure to UV light), and their antibacterial and morphological differences. Four effects of the Ag-complexes were evidenced by transmission electronic microscopy (TEM) and scanning electronic microscopy (SEM): DNA condensation, membrane disruption, shedding of cytoplasmic material and silver compound microcrystal penetration of bacteria. 5-Chlorosalicylic acid (5Cl) and sodium 4-aminosalicylate (4A) were the most effective ligands for synthesizing silver complexes with high levels of antibacterial activity. However, Ag-5Cl was the most stable against exposure UV light (365 nm). Cytotoxic effects were tested against two kinds of eukaryotic cells: murine fibroblast cells (T10 1/2) and human epithelial ovarian cancer cells (A2780). The main ob

The antinociceptive effect of resveratrol in bone cancer pain is inhibited by the Silent Information Regulator 1 inhibitor selisistat

© 2018 Royal Pharmaceutical SocietyObjectives: To study the antinociceptive effect of single and repeated doses of resveratrol in a bone cancer pain model, and whether this effect is prevented by the Silent Information Regulator 1 (SIRT1) inhibitor selisistat. Methods: The femoral intercondylar bone of BALB/c mice was injected with 1 000 000 BJ3Z cancer cells. Bone resorption and tumour mass growth (measured by in vivo X-ray and fluorescence imaging), as well as mechanical nociceptive thresholds (von Frey device) and dynamic functionality (rotarod machine), were evaluated during the following 4 weeks. Acute resveratrol (100 mg/kg i.p.) and/or selisistat (10 mg/kg s.c.) were administered on day 14. Chronic resveratrol (100 mg/kg i.p., daily) and/or selisistat (0.5 μg/h s.c., Alzet pump) were administered between days 14 and 20. Key findings: Tumour growth gradually incremented until day 31, while mechanical hyperalgesia started on day 3 after cancer cell injection. Acute resveratrol in

Peptide Targeted Gold Nanoplatform Carrying miR-145 Induces Antitumoral Effects in Ovarian Cancer Cells

One of the recent attractive therapeutic approaches for cancer treatment is restoring downregulated microRNAs. They play an essential muti-regulatory role in cellular processes such as proliferation, differentiation, survival, apoptosis, cell cycle, angiogenesis, and metastasis, among others. In this study, a gold nanoplatform (GNPF) carrying miR-145, a downregulated microRNA in many cancer types, including epithelial ovarian cancer, was designed and synthesized. For targeting purposes, the GNPF was functionalized with the FSH33 peptide, which provided selectivity for ovarian cancer, and loaded with the miR-145 to obtain the nanosystem GNPF-miR-145. The GNPF-mir-145 was selectively incorporated in A2780 and SKOV3 cells and significantly inhibited cell viability and migration and exhibited proliferative and anchor-independent growth capacities. Moreover, it diminished VEGF release and reduced the spheroid size of ovarian cancer through the damage of cell membranes, thus decreasing cell viability and possibly activating apoptosis. These results provide important advances in developing miR-based therapies using nanoparticles as selective vectors and provide approaches for in vivo evaluation.</jats:p