Publications by authors named "Manuel Arruebo"

87 Publications

Nondestructive production of exosomes loaded with ultrathin palladium nanosheets for targeted bio-orthogonal catalysis.

Nat Protoc 2021 01 27;16(1):131-163. Epub 2020 Nov 27.

Instituto de Nanociencia y Materiales de Aragón (INMA), CSIC-Universidad de Zaragoza, Zaragoza, Spain.

The use of exosomes as selective delivery vehicles of therapeutic agents, such as drugs or hyperthermia-capable nanoparticles, is being intensely investigated on account of their preferential tropism toward their parental cells. However, the methods used to introduce a therapeutic load inside exosomes often involve disruption of their membrane, which may jeopardize their targeting capabilities, attributed to their surface integrins. On the other hand, in recent years bio-orthogonal catalysis has emerged as a new tool with a myriad of potential applications in medicine. These bio-orthogonal processes, often based on Pd-catalyzed chemistry, would benefit from systems capable of delivering the catalyst to target cells. It is therefore highly attractive to combine the targeting capabilities of exosomes and the bio-orthogonal potential of Pd nanoparticles to create new therapeutic vectors. In this protocol, we provide detailed information on an efficient procedure to achieve a high load of catalytically active Pd nanosheets inside exosomes, without disrupting their membranes. The protocol involves a multistage process in which exosomes are first harvested, subjected to impregnation with a Pd salt precursor followed by a mild reduction process using gas-phase CO, which acts as both a reducing and growth-directing agent to produce the desired nanosheets. The technology is scalable, and the protocol can be conducted by any researcher having basic biology and chemistry skills in ~3 d.
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http://dx.doi.org/10.1038/s41596-020-00406-zDOI Listing
January 2021

Antimicrobial Wound Dressings against Fluorescent and Methicillin-Sensitive Intracellular Pathogenic Bacteria.

ACS Appl Mater Interfaces 2020 Nov 4;12(46):51302-51313. Epub 2020 Nov 4.

Department of Chemical Engineering, Aragon Institute of Nanoscience (INA), University of Zaragoza, Campus Río Ebro-Edificio I+D, C/Mariano Esquillor S/N, 50018 Zaragoza, Spain.

There is limited evidence indicating that drug-eluting dressings are clinically more effective than simple conventional dressings. To shed light on this concern, we have performed evidence-based research to evaluate the antimicrobial action of thymol (THY)-loaded antimicrobial dressings having antibiofilm forming ability, able to eradicate intracellular and extracellular pathogenic bacteria. We have used four different strains, including the ATCC 25923 strain, the Newman strain (methicillin-sensitive strain, MSSA) expressing the coral green fluorescent protein from the vector pCN47, and two clinical reference strains, Newman-(MSSA) and USA300-(methicillin-resistant strain), as traceable models of pathogenic bacteria commonly infecting skin and soft tissues. Compared to non-loaded dressings, THY-loaded polycaprolactone-based electrospun dressings were also able to eliminate pathogenic bacteria in coculture models based on infected murine macrophages. In addition, by using confocal microscopy and the conventional microdilution plating method, we corroborated the successful ability of THY in preventing also biofilm formation. Herein, we demonstrated that the use of wound dressings loaded with the natural monoterpenoid phenol derivative THY are able to eliminate biofilm formation and intracellular methicillin-sensitive more efficiently than with their corresponding THY-free counterparts.
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http://dx.doi.org/10.1021/acsami.0c17043DOI Listing
November 2020

Isolation of exosomes from whole blood by a new microfluidic device: proof of concept application in the diagnosis and monitoring of pancreatic cancer.

J Nanobiotechnology 2020 Oct 22;18(1):150. Epub 2020 Oct 22.

Department of Chemical Engineering, University of Zaragoza, 50018, Zaragoza, Spain.

Background: Exosomes are endocytic-extracellular vesicles with a diameter around 100 nm that play an essential role on the communication between cells. In fact, they have been proposed as candidates for the diagnosis and the monitoring of different pathologies (such as Parkinson, Alzheimer, diabetes, cardiac damage, infection diseases or cancer).

Results: In this study, magnetic nanoparticles (FeONPs) were successfully functionalized with an exosome-binding antibody (anti-CD9) to mediate the magnetic capture in a microdevice. This was carried out under flow in a 1.6 mm (outer diameter) microchannel whose wall was in contact with a set of NdFeB permanent magnets, giving a high magnetic field across the channel diameter that allowed exosome separation with a high yield. To show the usefulness of the method, the direct capture of exosomes from whole blood of patients with pancreatic cancer (PC) was performed, as a proof of concept. The captured exosomes were then subjected to analysis of CA19-9, a protein often used to monitor PC patients.

Conclusions: Here, we describe a new microfluidic device and the procedure for the isolation of exosomes from whole blood, without any need of previous isolation steps, thereby facilitating translation to the clinic. The results show that, for the cases analyzed, the evaluation of CA19-9 in exosomes was highly sensitive, compared to serum samples.
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http://dx.doi.org/10.1186/s12951-020-00701-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7579907PMC
October 2020

Controlling Particle Size and Release Kinetics in the Sustained Delivery of Oral Antibiotics Using pH-Independent Mucoadhesive Polymers.

Mol Pharm 2020 09 5;17(9):3314-3327. Epub 2020 Aug 5.

Department of Chemical Engineering, Aragon Institute of Nanoscience (INA), University of Zaragoza, Aragón Materials Science Institute, ICMA, Campus Río Ebro-Edificio I+D, C/ Poeta Mariano Esquillor S/N, 50018 Zaragoza, Spain.

Copolymers synthesized from acrylic acid and methacrylic acid used as gastroprotective and mucoadhesive enteric coatings have been used to prepare micro- (∼2 μm), submicro- (∼200 nm), and nanoparticles (∼20 nm) containing rifampicin (Rif) to obtain time-controlled drug release kinetics. Different particle sizes and drug release kinetics have been obtained using different synthesis conditions and fabrication techniques including the use of an electrosprayer and an interdigital microfabricated micromixer. The antimicrobial action of the encapsulated Rif has been demonstrated against ATCC 25923 and compared with the effect of the equivalent dose of the free macrolide antibiotic. At low concentrations, the encapsulated antibiotic showed superior antimicrobial activity than the free drug. The stability of the developed particles has been evaluated under simulated gastric and intestinal conditions. At the concentrations tested, a reduced cytotoxicity against different human cell lines was observed after analyzing their subcytotoxic doses and the influence on their cell cycle by flow cytometry. Drug release kinetics can be tuned by adjusting particle sizes, and it would be possible to reach the minimum inhibitory concentration or the minimum bactericidal concentration at different time points depending on the medical needs.
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http://dx.doi.org/10.1021/acs.molpharmaceut.0c00408DOI Listing
September 2020

Microflow Nanoprecipitation of Positively Charged Gastroresistant Polymer Nanoparticles of Eudragit RS100: A Study of Fluid Dynamics and Chemical Parameters.

Materials (Basel) 2020 Jun 30;13(13). Epub 2020 Jun 30.

Department of Chemical Engineering and Environmental Technology, Aragon Institute of Nanoscience (INA), Instituto de Ciencia de Materiales de Aragon-ICMA, University of Zaragoza, 50018 Zaragoza, Spain.

The objective of the present work was to produce gastroresistant Eudragit RS100 nanoparticles by a reproducible synthesis approach that ensured mono-disperse nanoparticles under the size of 100 nm. Batch and micromixing nanoprecipitation approaches were selected to produce the demanded nanoparticles, identifying the critical parameters affecting the synthesis process. To shed some light on the formulation of the targeted nanoparticles, the effects of particle size and homogeneity of fluid dynamics, and physicochemical parameters such as polymer concentration, type of solvent, ratio of solvent to antisolvent, and total flow rate were studied. The physicochemical characteristics of resulting nanoparticles were studied applying dynamic light scattering (DLS) particle size analysis and electron microscopy imaging. Nanoparticles produced using a micromixer demonstrated a narrower and more homogenous distribution than the ones obtained under similar conditions in conventional batch reactors. Besides, fluid dynamics ensured that the best mixing conditions were achieved at the highest flow rate. It was concluded that nucleation and growth events must also be considered to avoid uncontrolled nanoparticle growth and evolution at the collection vial. Further, rifampicin-encapsulated nanoparticles were prepared using both approaches, demonstrating that the micromixing-assisted approach provided an excellent control of the particle size and polydispersity index. Not only the micromixing-assisted nanoprecipitation promoted a remarkable control in the nanoparticle formulation, but also it enhanced drug encapsulation efficiency and loading, as well as productivity. To the best of our knowledge, this was the very first time that drug-loaded Eudragit RS100 nanoparticles (NPs) were produced in a continuous fashion under 100 nm (16.5 ± 4.3 nm) using microreactor technology. Furthermore, we performed a detailed analysis of the influence of various fluid dynamics and physicochemical parameters on the size and uniformity of the resulting nanoparticles. According to these findings, the proposed methodology can be a useful approach to synthesize a myriad of nanoparticles of alternative polymers.
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http://dx.doi.org/10.3390/ma13132925DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7372341PMC
June 2020

Drug-eluting wound dressings having sustained release of antimicrobial compounds.

Eur J Pharm Biopharm 2020 Jul 28;152:327-339. Epub 2020 May 28.

Department of Chemical Engineering, Aragon Institute of Nanoscience (INA), University of Zaragoza, Campus Río Ebro-Edificio I+D, C/ Poeta Mariano Esquillor S/N, 50018 Zaragoza, Spain; Networking Research Center on Bioengineering, Biomaterials and Nanomedicine, CIBER-BBN, 28029 Madrid, Spain; Aragon Health Research Institute (IIS Aragon), 50009 Zaragoza, Spain.

Wound healing is a complex and costly public health problem that should be timely addressed to achieve a rapid and adequate tissue repair avoiding or even eliminating potential pathogenic infection. Chronic infected non-healing wounds represent a serious concern for health care systems. Efficient wound dressings with tailored therapy having the best response and highest safety margin for the management of chronic non-healing wounds are still needed. The use of novel wound dressing materials has emerged as a promising tool to fulfil these requirements. In this work, asymmetric electrospun polycaprolactone (PCL)-based nanofibers (NFs) were decorated with electrosprayed poly(lactic-co-glycolic acid) microparticles (PLGA MPs) containing the natural antibacterial compound thymol (THY) in order to obtain drug eluting antimicrobial dressings having sustained release. The synthesized dressings successfully inhibited the in vitro growth of Staphylococcus aureus ATCC 25923, showing also at the same doses cytocompatibility on human dermal fibroblasts and keratinocyte cultures after treatment for 24 h, which was not observed when using free thymol. An in vivo murine excisional wound splinting model, followed by the experimental infection of the wounds with S. aureus and their treatment with the synthesized dressings, pointed to the reduction of the bacterial load in wounds after 7 days, though the total elimination of the infection was not reached. The findings indicated the relevance of the direct contact between the dressings and the bacteria, highlighting the need to tune their design considering the wound surface and the nature of the antimicrobial cargo contained.
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http://dx.doi.org/10.1016/j.ejpb.2020.05.025DOI Listing
July 2020

Novel intracellular antibiotic delivery system against : cloxacillin-loaded poly(d,l-lactide-co-glycolide) acid nanoparticles.

Nanomedicine (Lond) 2020 05 6;15(12):1189-1203. Epub 2020 May 6.

Microbiology Department, Hospital Universitari Germans Trias i Pujol, Fundació Institut d'Investigació en Ciències de la Salut GermansTrias i Pujol, Universitat Autònoma de Barcelona, Badalona, Spain.

First, to compare minimum inhibitory concentrations (MIC) of free cloxacillin and cloxacillin-containing nanoparticles (NP) against methicillin-susceptible (MSSA) and resistant (MRSA) and second, to assess NP antimicrobial activity against intracellular . Poly(d,l-lactide-co-glycolide) acid (PLGA)-NP were loaded with cloxacillin and physico-chemically characterized. MICs were determined for reference strains Newman-(MSSA) and USA300-(MRSA). Murine alveolar macrophages were infected, and bacterial intracellular survival was assessed after incubating with free-cloxacillin or PLGA-cloxacillin-NP. For both isolates, MICs for antibiotic-loaded-NP were lower than those obtained with free cloxacillin, indicating that the drug encapsulation improves antimicrobial activity. A sustained antibiotic release was demonstrated when using the PLGA-cloxacillin-NP. When considering the lowest concentrations, the use of drug-loaded NP enabled a higher reduction of intracellular bacterial load.
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http://dx.doi.org/10.2217/nnm-2019-0371DOI Listing
May 2020

Antibacterial Effect of Thymol Loaded SBA-15 Nanorods Incorporated in PCL Electrospun Fibers.

Nanomaterials (Basel) 2020 Mar 27;10(4). Epub 2020 Mar 27.

Department of Chemical Engineering. Aragon Institute of Nanoscience (INA), University of Zaragoza, Campus Río Ebro-Edificio I+D, C/Poeta Mariano Esquillor S/N, 50018 Zaragoza, Spain.

For the effective management of infected chronic wounds, the incorporation of antimicrobial drugs into wound dressings can increase their local availability at the infection site. Mesoporous silicon dioxide SBA-15 is an excellent drug carrier with tunable drug release kinetics. In this work, synthesized SBA-15 loaded with the natural antimicrobial compound thymol (THY) was incorporated into polycaprolactone (PCL) electrospun nanofibers to obtain an advanced wound dressing. Rod-shaped particles with internal parallel channels oriented along the longitudinal axis (diameter: 138 ± 30 nm, length: 563 ± 100 nm) were loaded with 70.8 wt.% of THY. Fiber mats were prepared using these particles as nanofillers within polycaprolactone (PCL) electrospun fibers. The resulting mats contained 5.6 wt.% of THY and more than half of this loading was released in the first 7 h. This release would prevent an initial bacterial colonization and also inhibit or eliminate bacterial growth as in vitro shown against ATCC 25923. Minimal inhibitory concentration (MIC: 0.07 mg/mL) and minimal bactericidal concentration (MBC: 0.11 mg/mL) of released THY were lower than the amount of free THY required, demonstrating the benefit of drug encapsulation for a more efficient bactericidal capacity due to the direct contact between mats and bacteria.
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http://dx.doi.org/10.3390/nano10040616DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7221837PMC
March 2020

Customized hybrid and NIR-light triggered thermoresponsive drug delivery microparticles synthetized by photopolymerization in a one-step flow focusing continuous microreactor.

Colloids Surf B Biointerfaces 2020 Jun 25;190:110904. Epub 2020 Feb 25.

Department of Chemical Engineering. Aragon Institute of Nanoscience (INA) and Instituto de Ciencia de Materiales de Aragón (ICMA), Universidad de Zaragoza-CSIC, University of Zaragoza, Campus Río Ebro-Edificio I+D, C/ Poeta Mariano Esquillor S/N, 50018, Zaragoza, Spain; Aragon Health Research Institute (IIS Aragón), 50009, Zaragoza, Spain; Networking Research Center on Bioengineering, Biomaterials and Nanomedicine, CIBER-BBN, 28029, Madrid, Spain.

Photopolymerization is a selective technique that takes advantage of light-sensitive molecules to initiate and propagate monomeric structures to render covalently bonded macromolecular materials structures known as polymers. Herein, we present a novel one-step microfluidic synthesis of customized hybrid-thermoresponsive Poly(N-isopropylacrylamide) (PNIPAm) based microparticles (MPs) containing plasmonic hollow gold nanoparticles (HGNPs) and bupivacaine (BVP) used as a model drug. Those hybrid microparticles were prepared using a flow-focusing microreactor coupled to a UV LED device built with a simple outer PTFE tubing and an inner flexible capillary. Different tubing characteristics and flow rate ratios were altered in order to control the size of the resulting microparticles. In addition, components such as monomer, crosslinker and photoinitiator concentrations, as well as LED intensity and irradiation time were tuned to obtain different MPs and their characteristics and polymerization rates were compared by Gel Permeation Chromatography (GPC). Thermoresponsive properties were analyzed and the presence of HGNPs was confirmed in light-activated triggered drug release applications. Bupivacaine loading and release studies were evaluated with the resulting hollow and solid microparticles (which were obtained depending on the polymerization rate used) and their temperature responsiveness was assessed using a NIR laser when HGNPs were present in the constructs. Finally, cytotoxicity studies, cell-cycle arrest and apoptotic induction were carried out to certify their suitability for further biomedical applications to be used as triggerable drug depots.
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http://dx.doi.org/10.1016/j.colsurfb.2020.110904DOI Listing
June 2020

Metallocenyl 7-ACA Conjugates: Antibacterial Activity Studies and Atomic-Resolution X-ray Crystal Structure with CTX-M β-Lactamase.

Chembiochem 2020 08 16;21(15):2187-2195. Epub 2020 Apr 16.

Department of Organic Chemistry, Faculty of Chemistry, University of Łódź, Tamka 12, 91-403, Łódź, Poland.

The conjugation of organometallic groups to current β-lactam antibiotics is a field of increasing study due to the ability of certain organometallic groups to enhance the antibiotic potency of these drugs. Herein, we report the antibacterial properties of two metallocenyl (ferrocenyl and ruthenocenyl) 7-aminocephalosporanic acid (7-ACA) antibiotic conjugates. Continuing a trend we found in our previous studies, the ruthenocenyl conjugate showed greater antibacterial activity than its ferrocenyl counterpart. Compared with the previously published 7-aminodesacetoxycephalosporanic acid (7-ADCA) conjugates, the 3-acetyloxymethyl group significantly improved the compounds' activity. Furthermore, the Rc-7-ACA compound was more active against clinical Staphylococcus aureus isolates than the ampicillin reference. Noticeably, neither of the two new compounds showed an undesirable toxic effect in HeLa and L929 cells at the concentrations at which they displayed strong antibacterial effects. The antibacterial activity of the two metallocenyl 7-ACA derivatives was further confirmed by scanning electron microscopy (SEM). SEM micrographs showed that bacteria treated with metallocenyl 7-ACA derivatives feature cell wall damage and morphology changes. Using a CTX-M-14 β-lactamase competition assay based on nitrocefin hydrolysis, we showed that the Rc-7-ACA bound more favorably to CTX-M-14 than its ferrocenyl counterpart, again confirming the superiority of the ruthenocenyl moiety over the ferrocenyl one in interacting with proteins. We also report a 1.47 Å resolution crystal structure of Rc-7-ACA in complex with the CTX-M-14 E166A mutant, an enzyme sharing a similar active site configuration with penicillin-binding proteins, the molecular target of β-lactam antibiotics. These results strengthen the case for the antibacterial utility of the Rc and Fc groups.
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http://dx.doi.org/10.1002/cbic.202000054DOI Listing
August 2020

Local delivery of bone morphogenetic protein-2 from near infrared-responsive hydrogels for bone tissue regeneration.

Biomaterials 2020 05 21;241:119909. Epub 2020 Feb 21.

Hospital Universitario La Paz-IdiPAZ, Paseo de La Castellana 261, Madrid, 28046, Spain; CIBER de Bioingenieria, Biomateriales y Nanomedicina, CIBER-BBN, Spain. Electronic address:

Achievement of spatiotemporal control of growth factors production remains a main goal in tissue engineering. In the present work, we combined inducible transgene expression and near infrared (NIR)-responsive hydrogels technologies to develop a therapeutic platform for bone regeneration. A heat-activated and dimerizer-dependent transgene expression system was incorporated into mesenchymal stem cells to conditionally control the production of bone morphogenetic protein 2 (BMP-2). Genetically engineered cells were entrapped in hydrogels based on fibrin and plasmonic gold nanoparticles that transduced incident energy of an NIR laser into heat. In the presence of dimerizer, photoinduced mild hyperthermia induced the release of bioactive BMP-2 from NIR-responsive cell constructs. A critical size bone defect, created in calvaria of immunocompetent mice, was filled with NIR-responsive hydrogels entrapping cells that expressed BMP-2 under the control of the heat-activated and dimerizer-dependent gene circuit. In animals that were treated with dimerizer, NIR irradiation of implants induced BMP-2 production in the bone lesion. Induction of NIR-responsive cell constructs conditionally expressing BMP-2 in bone defects resulted in the formation of new mineralized tissue, thus indicating the therapeutic potential of the technological platform.
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http://dx.doi.org/10.1016/j.biomaterials.2020.119909DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7263445PMC
May 2020

Light-triggered nanoparticles for pain management.

Expert Opin Drug Deliv 2020 05 5;17(5):627-633. Epub 2020 Mar 5.

Department of Chemical Engineering and Environmental Technologies, Aragon Nanoscience Institute, University of Zaragoza, Zaragoza, Spain.

: Recent technological progress in pain management includes patient´s stratification depending on their disease subtype, prognosis, risk, or treatment response using data analysis and genetic testing in order to select the most appropriate drug for each group. A spatiotemporal control on the release of the selected anesthetic drug is also desirable in order to minimize side effects and to provide the patient with the appropriate dose above the therapeutic threshold and below the maximum desirable concentration. Light can be used non-invasively as an exogenous trigger to allow multiple drug administrations with precise spatiotemporal control. By controlling light fluence/irradiance, pulse structure, and duration of the irradiation drug release kinetics can be controlled in a pulsatile manner to release totally or partially the drug loaded into particulate carriers.: Recent advances in the field of light-triggered nanoparticles used in pain management specially those studies which include preclinical models are reviewed.: Two decades later after the first light-sensitive drug delivery systems reported still several limitations hinder their clinical translation. Additional efforts should be undertaken to understand the nanoparticles biological fate, to satisfy their large-scale production, and to facilitate the technology to apply this therapeutic approach at a low cost.
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http://dx.doi.org/10.1080/17425247.2020.1737670DOI Listing
May 2020

Electrospun anti-inflammatory patch loaded with essential oils for wound healing.

Int J Pharm 2020 Mar 22;577:119067. Epub 2020 Jan 22.

Regenerative Medicine Program, Houston Methodist Research Institute, Houston, TX, USA; Orthopedics & Sports Medicine, Houston Methodist Hospital, Houston, TX, USA.

Wound healing is a complex process that consists of three overlapping phases: inflammation, proliferation, and remodeling. A bacterial infection can increase inflammation and delay this process. Microorganisms are closely related to the innate immune system, such as macrophages and neutrophils, as they can start an inflammatory cascade. Essential oils play an important role in the inhibition and prevention of bacterial growth due to their ability to reduce antimicrobial resistance. The possibility to find a strategy that combines antimicrobial and anti-inflammatory properties is particularly appealing for wound healing. In this work, we showcase a variety of patches based on electrospun polycaprolactone (PCL) nanofibers loaded with natural compounds derived from essential oils, such as thymol (THY) and tyrosol (TYR), to achieve reduced inflammation. In addition, we compared the effect these essential oils have on activated macrophages when incorporated into the PCL patch. Specifically, we demonstrate that PCL-THY resulted in more efficient down-regulation of pro-inflammatory genes related to the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κb) pathway when compared to PCL-TYR and the combination patch containing TYR and THY (i.e., PCL-TYR-THY). Furthermore, PCL-THY displayed low affinity for cell attachment, which may hinder wound adherence and integration. Overall, our results indicate that THY-loaded patches could serve as promising candidates for the fabrication of dressings that incorporate bactericidal and anti-inflammatory properties while simultaneously avoiding the limitations of traditional antibiotic-loaded devices.
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http://dx.doi.org/10.1016/j.ijpharm.2020.119067DOI Listing
March 2020

Organometallic ciprofloxacin conjugates with dual action: synthesis, characterization, and antimicrobial and cytotoxicity studies.

Dalton Trans 2020 Feb 18;49(5):1403-1415. Epub 2019 Dec 18.

Faculty of Chemistry, Department of Organic Chemistry, University of Łódź, Tamka 12, 91-403 Łódź, Poland.

The synthesis, characterization and biological activity of six bioorganometallic conjugates of ciprofloxacin with ferrocenyl, ruthenocenyl and cymantrenyl entities are described. Their antimicrobial activities were investigated against Gram-positive bacteria, Gram-negative bacteria and bloodstream forms of Trypanosoma brucei. Furthermore, the morphological changes of bacterial cells upon treatment with the conjugates were examined by scanning electron microscopy. In addition, the cytotoxicity of the conjugates against tumor and normal mammalian cells was also investigated. The results showed that conjugation of an organometallic moiety can significantly enhance the antimicrobial activity of the antibiotic ciprofloxacin drug. It was found that N-alkyl cymantrenyl and ruthenocenyl ciprofloxacin conjugates were the most effective derivatives although other conjugates also showed significant antimicrobial activity. The increase in the antimicrobial activity was most likely due to two independent mechanisms of action. The first mechanism is due to the bacterial topoisomerase inhibitory activity of ciprofloxacin while the second mechanism can be attributed to the generation of reactive oxygen species caused by the organometallic moiety. The presence of two modes of action enables the conjugates to kill bacteria in their stationary growth phase and to overcome the drug resistance of S. aureus strains. In addition, the conjugates showed promising selectivity toward bacterial and parasitic cells over mammalian cells.
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http://dx.doi.org/10.1039/c9dt03948aDOI Listing
February 2020

Extracellular Vesicles-Based Biomarkers Represent a Promising Liquid Biopsy in Endometrial Cancer.

Cancers (Basel) 2019 Dec 12;11(12). Epub 2019 Dec 12.

Translational Medical Oncology Group (Oncomet), Health Research Institute of Santiago de Compostela (IDIS), University Hospital of Santiago de Compostela (SERGAS), Trav. Choupana s/n, 15706 Santiago de Compostela, Spain.

Tumor-derived extracellular vesicles (EVs) are secreted in large amounts into biological fluids of cancer patients. The analysis of EVs cargoes has been associated with patient´s outcome and response to therapy. However, current technologies for EVs isolation are tedious and low cost-efficient for routine clinical implementation. To explore the clinical value of circulating EVs analysis we attempted a proof-of-concept in endometrial cancer (EC) with ExoGAG, an easy to use and highly efficient new technology to enrich EVs. Technical performance was first evaluated using EVs secreted by Hec1A cells. Then, the clinical value of this strategy was questioned by analyzing the levels of two well-known tissue biomarkers in EC, L1 cell adhesion molecule (L1CAM) and Annexin A2 (ANXA2), in EVs purified from plasma in a cohort of 41 EC patients and 20 healthy controls. The results demonstrated the specific content of ANXA2 in the purified EVs fraction, with an accurate sensitivity and specificity for EC diagnosis. Importantly, high ANXA2 levels in circulating EVs were associated with high risk of recurrence and non-endometrioid histology suggesting a potential value as a prognostic biomarker in EC. These results also confirmed ExoGAG technology as a robust technique for the clinical implementation of circulating EVs analyses.
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http://dx.doi.org/10.3390/cancers11122000DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6966595PMC
December 2019

Triggered drug release from hybrid thermoresponsive nanoparticles using near infrared light.

Nanomedicine (Lond) 2020 02 27;15(3):219-234. Epub 2019 Nov 27.

Department of Chemical Engineering, Aragon Institute of Nanoscience (INA), University of Zaragoza, Campus Río Ebro-Edificio I+D, C/ Poeta Mariano Esquillor S/N, 50018-Zaragoza, Spain.

Developing hybrid poly(N-isopropylacrylamide)-based nanogels decorated with plasmonic hollow gold nanoparticles for on-demand drug delivery and their physico-chemical characterization, bupivacaine loading and release ability upon light irradiation, and cell viability. Hollow gold nanoparticles were prepared by galvanic replacement reaction; poly(N-isopropylacrylamide)-based nanogels were synthesized via precipitation polymerization and their electrostatic coupling was accomplished using poly(allylamine hydrochloride) as cationic polyelectrolyte linker. Colloidal stability of the resulted hybrid nanovectors was demonstrated under physiological conditions together with their fast response and excellent heating efficiency after light stimulation, indicating their potential use as triggered drug-delivery vectors. Moreover, their influence on cell metabolism and cell cycle under subcytotoxic doses were studied showing excellent cytocompatibility.
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http://dx.doi.org/10.2217/nnm-2019-0270DOI Listing
February 2020

Antimicrobial Electrospun Polycaprolactone-Based Wound Dressings: An Study About the Importance of the Direct Contact to Elicit Bactericidal Activity.

Adv Wound Care (New Rochelle) 2019 Sep 9;8(9):438-451. Epub 2019 Aug 9.

Department of Chemical Engineering, Aragon Institute of Nanoscience (INA), University of Zaragoza, Zaragoza, Spain.

To prepare efficient antibacterial carvacrol (CAR) and thymol (THY)-loaded electrospun polycaprolactone (PCL)-based wound dressings. Using electrospinning we were able to prepare wound dressings with antimicrobial action thanks to their large surface per volume ratio, which allows their loading with therapeutic amounts of active principles. By nuclear magnetic resonance we demonstrated that the antimicrobial compounds are donors of hydrogen bonds to the ester functional group in PCL, which acts as acceptor and that intermolecular interaction is responsible for the high drug loading achieved. Those mats loaded with CAR and THY without the use of solubilizing agents were able to completely eradicate both Gram-positive ( ATCC 25923) and Gram-negative ( S17 strain) bacteria at doses inferior to the ones needed when using the free nonsupported compounds. A superior antimicrobial action was observed for THY and CAR against Gram-negative bacteria than against Gram-positive bacteria, despite the higher hydrophilicity of the outer layer of Gram-negative bacteria. We demonstrate that a direct contact between the bacteria and the dressing is required to elicit antimicrobial action. We also evaluated drug loadings by gas chromatography coupled with mass spectrometry and nuclear magnetic resonance validating a new analytical approach. Finally we were able to visualize the pathogenic bacteria on the dressings by confocal microscopy. The interaction between the PCL-based mat and the pathogenic bacteria is a key issue to achieve complete pathogen eradication. Under no-contact conditions, released CAR or THY from the electrospun mats did not exert any antimicrobial action at the doses tested.
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http://dx.doi.org/10.1089/wound.2018.0893DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6855275PMC
September 2019

Reversible stimuli-responsive nanomaterials with on-off switching ability for biomedical applications.

J Control Release 2019 11 20;314:162-176. Epub 2019 Oct 20.

Department of Chemical Engineering and Environmental Technologies. Aragon Nanoscience Institute. University of Zaragoza, Campus Río Ebro-Edificio I+D, C/Poeta Mariano Esquillor S/N, 50018, Zaragoza, Spain; Aragon Health Research Institute (IIS Aragon), 50009-Zaragoza, Spain; Networking Research Center on Bioengineering, Biomaterials and Nanomedicine, CIBER-BBN, 28029, Madrid, Spain.

Many long-acting extended drug release systems can provide controlled or sustained release of therapeutic payloads. In the majority of those systems drug release cannot be stopped once it has started because they operate autonomously regardless of the evolution of the treatment and/or the patient´s needs. However, in several pathologies such as diabetes, hormonal disorders, pain management, etc. a pulsatile drug release is required to adjust the dose of drug release to the specific needs in a spatio-temporal manner. Additionally, in other pathologies such as cancer or antimicrobial therapy the release of the drug with spatio-temporal control to prevent unwanted side effects represents an unmet need. With this aim reversible stimuli-responsive nanomaterials with an on-off switching ability have been developed in order to provide a spatio-temporal control of the drug released. Those systems can be activated in response to exogenous (light, magnetic field, electrical fields, etc.) or endogenous triggers (pH, enzyme-substrate complex formation, protein-cell binding, etc.) thanks to the use of reversible phase-transition materials. In this review we compile in vitro and preclinical results in which those materials have been successfully used. The types of stimuli used to trigger drug release as well as the different nanomaterials used are reviewed in order to provide a general overview of the field. We anticipate that further studies in this field will be expanded towards the development of multimodal hybrid systems which combine therapy and imaging while reporting the evolution of the treatment in real time.
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http://dx.doi.org/10.1016/j.jconrel.2019.10.036DOI Listing
November 2019

Cancer-derived exosomes loaded with ultrathin palladium nanosheets for targeted bioorthogonal catalysis.

Nat Catal 2019 Oct 9;2(10):864-872. Epub 2019 Sep 9.

Cancer Research UK Edinburgh Centre, MRC Institute of Genetics & Molecular Medicine, University of Edinburgh, Crewe Road South, Edinburgh EH4 2XR, UK.

The transformational impact of bioorthogonal chemistries has inspired new strategies for the synthesis of bioactive agents through non-natural means. Among these, palladium (Pd) catalysts have played a prominent role in the growing subfield of bioorthogonal catalysis by producing xenobiotics and uncaging biomolecules in living systems. However, delivering catalysts selectively to specific cell types still lags behind catalyst development. Here we have developed a bio-artificial device consisting of cancer-derived exosomes loaded with Pd catalysts by a method that enables the controlled assembly of Pd nanosheets directly inside the vesicles. This hybrid system mediates Pd-triggered dealkylation reactions and inside cells and displays preferential tropism for their progenitor cells. The use of Trojan exosomes to deliver abiotic catalysts into designated cancer cells creates the opportunity for a new targeted therapy modality: exosome-directed catalyst prodrug therapy, whose first steps are presented herein with the cell-specific release of the anticancer drug panobinostat.
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http://dx.doi.org/10.1038/s41929-019-0333-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6795537PMC
October 2019

Targeted Release of Probiotics from Enteric Microparticulated Formulations.

Polymers (Basel) 2019 Oct 13;11(10). Epub 2019 Oct 13.

Department of Chemical Engineering. Aragón Institute of Nanoscience (INA), University of Zaragoza, Campus Río Ebro-Edificio I+D, C/ Poeta Mariano Esquillor S/N, 50018-Zaragoza, Spain.

The development of advanced probiotic delivery systems, which preserve bacteria from degradation of the gastrointestinal tract and achieve a targeted release mediated by pH-independent swelling, is of great interest to improve the efficient delivery of probiotic bacteria to the target tissue. Gram-positive and Gram-negative bacteria models ( (Moro) Hansen and Mocquot (ATCC® 4356™) and S17, respectively) have been successfully encapsulated for the first time in pH-independent microparticulate polymethacrylates (i.e., Eudraguard biotic) used for the targeted delivery of nutraceuticals to the colon. These bacteria have also been encapsulated within the mucoadhesive polymethacrylate Eudragit RS 100 widely used as targeted release formulation for active pharmaceutical ingredients. The enteric microparticles remained unaltered under simulated gastric conditions and released the contained viable microbial cargo under simulated intestinal conditions. Buoyancies of 90.2% and 57.3% for Eudragit and Eudraguard microparticles, respectively, and long-term stability (5 months) for the encapsulated microorganisms were found. Cytotoxicity of the microparticles formulated with both polymers was evaluated (0.5-20 mg/mL) on Caco-2 cells, showing high cytocompatibility. These results underline the suitability of the synthesized materials for the successful delivery of probiotic formulations to the target organ, highlighting for the first time the potential use of Eudraguard biotic as an effective enteric coating for the targeted delivery of probiotics.
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http://dx.doi.org/10.3390/polym11101668DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6835770PMC
October 2019

Efficient encapsulation of theranostic nanoparticles in cell-derived exosomes: leveraging the exosomal biogenesis pathway to obtain hollow gold nanoparticle-hybrids.

Nanoscale 2019 Oct 9;11(40):18825-18836. Epub 2019 Oct 9.

Networking Research Center of Bioengineering, Biomaterials and Nanomedicine, CIBER-BBN, 28029-Madrid, Spain and Instituto Aragonés de Ciencias de la Salud (IACS), Centro de Investigación Biomédica de Aragón (CIBA), 50009-Zaragoza, Spain and IIS Aragón(IISA), Centro de Investigación Biomédica de Aragón (CIBA), 50009-Zaragoza, Spain and Fundación ARAID. Avda. Ranillas, 1-D, planta 2ª, oficina b, 50018-Zaragoza, Spain.

Exosomes can be considered natural targeted delivery systems able to carry exogenous payloads, drugs or theranostic nanoparticles (NPs). This work aims to combine the therapeutic capabilities of hollow gold nanoparticles (HGNs) with the unique tumor targeting properties provided by exosomes. Here, we tested different methods to encapsulate HGNs (capable of absorbing light in the NIR region for selective thermal ablation) into murine melanoma cells derived exosomes (B16-F10-exos), including electroporation, passive loading by diffusion, thermal shock, sonication and saponin-assisted loading. These methods gave less than satisfactory results: although internalization of relatively large NPs into B16-F10-exos was achieved by almost all the physicochemical methods tested, only about 15% of the exosomes were loaded with NPs and several of those processes had a negative effect regarding the morphology and integrity of the loaded exosomes. In a different approach, B16-F10 cells were pre-incubated with PEGylated HGNs (PEG-HGNs) in an attempt to incorporate the NPs into the exosomal biogenesis pathway. The results were highly successful: exosomes recovered from the supernatant of the cell culture showed up to 50% of HGNs internalization. The obtained hybrid HGN-exosome vectors were characterized with a battery of techniques to make sure that internalization of HGNs did not affect exosome characteristics compared with other strategies. PEG-HGNs were released through the endosomal-exosome biogenesis pathway confirming that the isolated vesicles were exosomes.
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http://dx.doi.org/10.1039/c9nr06183eDOI Listing
October 2019

Gold nanoparticles for the in situ polymerization of near-infrared responsive hydrogels based on fibrin.

Acta Biomater 2019 12 27;100:306-315. Epub 2019 Sep 27.

Hospital Universitario La Paz-IdiPAZ, Paseo de la Castellana 261, Madrid 28046 Spain; CIBER de Bioingenieria, Biomateriales y Nanomedicina, CIBER-BBN, Spain. Electronic address:

Non-invasiveness and relative safety of photothermal therapy, which enables local hyperthermia of target tissues using a near infrared (NIR) laser, has attracted increasing interest. Due to their biocompatibility, amenability of synthesis and functionalization, gold nanoparticles have been investigated as therapeutic photothermal agents. In this work, hollow gold nanoparticles (HGNP) were coated with poly-l-lysine through the use of COOH-Poly(ethylene glycol)-SH as a covalent linker. The functionalized HGNP, which peak their surface plasmon resonance at 800 nm, can bind thrombin. Thrombin-conjugated HGNP conduct in situ fibrin polymerization, facilitating the process of generating photothermal matrices. Interestingly, the metallic core of thrombin-loaded HGNP fragmentates at physiological temperature. During polymerization process, matrices prepared with thrombin-loaded HGNP were loaded with genetically-modified stem cells that harbour a heat-activated and ligand-dependent gene switch for regulating transgene expression. NIR laser irradiation of resulting cell constructs in the presence of ligand successfully triggered transgene expression in vitro and in vivo. STATEMENT OF SIGNIFICANCE: Current technological development allows synthesis of gold nanoparticles (GNP) in a wide range of shapes and sizes, consistently and at scale. GNP, stable and easily functionalized, show low cytotoxicity and high biocompatibility. Allied to that, GNP present optoelectronic properties that have been exploited in a range of biomedical applications. Following a layer-by-layer functionalization approach, we prepared hollow GNP coated with a positively charged copolymer that enabled thrombin conjugation. The resulting nanomaterial efficiently catalyzed the formation of fibrin hydrogels which convert energy of the near infrared (NIR) into heat. The resulting NIR-responsive hydrogels can function as scaffolding for cells capable of controlled gene expression triggered by optical hyperthermia, thus allowing the deployment of therapeutic gene products in desired spatiotemporal frameworks.
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http://dx.doi.org/10.1016/j.actbio.2019.09.040DOI Listing
December 2019

Differences in levan nanoparticles depending on their synthesis route: Microbial vs cell-free systems.

Int J Biol Macromol 2019 Sep 27;137:62-68. Epub 2019 Jun 27.

Department of Chemical Engineering, University of Salamanca, Plaza Los Caídos s/n, Salamanca, Spain; Institute for Biomedical Research of Salamanca (IBSAL), Paseo de San Vicente, 58-182, Salamanca, Spain. Electronic address:

Differences between the levan obtained from bacteria and from cell-free systems were studied in this work. Results showed that both polymers are non-porous solids (type II isotherm with 20 m/g) with a main thermal decomposition at 200 °C and a negligible value of protein adsorption. Microbial levan produced nanoparticles of 90 nm in diameter whereas nanoparticles of 110 nm were obtained with the polymer obtained from a cell-free system. Both polymers behave as aggregates depending on the critical aggregation concentration. At the same time, that concentration depends on the technique used for the polymer synthesis. Cell-free system aggregation concentration is 0.24 mg/mL whereas a concentration of 0.05 mg/mL was found for the microbial system. In both cases, the average molecular weight of the aggregate is higher than 2000 kDa. These results highlight the existence of aggregation equilibrium for both polymers that has to be taken into account for future applications.
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http://dx.doi.org/10.1016/j.ijbiomac.2019.06.128DOI Listing
September 2019

Preparation and Identification of Optimal Synthesis Conditions for a Novel Alkaline Anion-Exchange Membrane.

Polymers (Basel) 2018 Aug 13;10(8). Epub 2018 Aug 13.

Department of Chemical and Biomolecular Engineering, Universidad de Cantabria, 39005 Santander, Spain.

The physicochemical and mechanical properties of new alkaline anion-exchange membranes (AAEMs) based on chitosan (CS) and poly(vinyl alcohol) (PVA) polymers doped with unsupported copper nanoparticles (NPs) and copper exchanged over different porous materials were investigated regarding ion-exchange capacity (IEC), OH conductivity, water uptake (WU), water vapor permeability (WVP), and thermal and mechanical resistance. The influence of the type of filler included in different morphologies and filler loading has been explored using copper exchanged materials such as the layered porous titanosilicate AM-4, layered stannosilicate UZAR-S3, and zeolites Y, MOR, and BEA. Compared to commercially available anion-exchange membranes, the best performing membranes in terms of WU, IEC, OH conductivity and WVP in this study were those containing 10 wt % of Cu-AM-4 and Cu-UZAR-S3, although 10 wt % Cu-MOR provided better mechanical strength at close values of WVP and anion conductivity. It was also observed that when Cu was exchanged in a porous silicate matrix, its oxidation state was lower than when embedded as unsupported metal NPs. In addition, the statistical analysis of variance determined that the electrochemical properties of the membranes were noticeably affected by both the type and filler loading, and influenced also by the copper oxidation state and content in the membrane, but their hydrophilic properties were more affected by the polymers. The largest significant effects were noticed on the water sorption and transport properties, which gives scope for the design of AAEMs for electrochemical and water treatment applications.
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http://dx.doi.org/10.3390/polym10080913DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6403638PMC
August 2018

Matryoshka-type gastro-resistant microparticles for the oral treatment of Mycobacterium tuberculosis.

Nanomedicine (Lond) 2019 Mar 8;14(6):707-726. Epub 2019 Feb 8.

Department of Chemical Engineering. Aragon Institute of Nanoscience (INA), University of Zaragoza, Campus Río Ebro-Edificio I+D, C/Poeta Mariano Esquillor S/N, Zaragoza 50018, Spain.

Aim: Production of Matryoshka-type gastroresistant microparticles containing antibiotic-loaded poly lactic-co-glycolic acid (PLGA) nanoparticles (NP) against Mycobacterium tuberculosis.

Materials & Methods: The emulsification and evaporation methods were followed for the synthesis of PLGA-NPs and methacrylic acid-ethyl acrylate-based coatings to protect rifampicin from degradation under simulated gastric conditions.

Results & Conclusion: The inner antibiotic-loaded NPs here reported can be released under simulated intestinal conditions whereas their coating protects them from degradation under simulated gastric conditions. The encapsulation does not hinder the antituberculosis action of the encapsulated antibiotic rifampicin. A sustained antibiotic release could be obtained when using the drug-loaded encapsulated NPs. Compared with the administration of the free drug, a more effective elimination of M. tuberculosis was observed when applying the NPs against infected macrophages. The antibiotic-loaded PLGA-NPs were also able to cross an in vitro model of intestinal barrier.
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http://dx.doi.org/10.2217/nnm-2018-0258DOI Listing
March 2019

Liver Expression of a MiniATP7B Gene Results in Long-Term Restoration of Copper Homeostasis in a Wilson Disease Model in Mice.

Hepatology 2019 07 20;70(1):108-126. Epub 2019 Mar 20.

Gene Therapy and Regulation of Gene Expression Program, CIMA, FIMA, University of Navarra, Navarra Institute for Health Research (IdisNA), Pamplona, Spain.

Gene therapy with an adeno-associated vector (AAV) serotype 8 encoding the human ATPase copper-transporting beta polypeptide (ATP7B) complementary DNA (cDNA; AAV8-ATP7B) is able to provide long-term copper metabolism correction in 6-week-old male Wilson disease (WD) mice. However, the size of the genome (5.2 kilobases [kb]) surpasses the optimal packaging capacity of the vector, which resulted in low-yield production; in addition, further analyses in WD female mice and in animals with a more advanced disease revealed reduced therapeutic efficacy, as compared to younger males. To improve efficacy of the treatment, an optimized shorter AAV vector was generated, in which four out of six metal-binding domains (MBDs) were deleted from the ATP7B coding sequence, giving rise to the miniATP7B protein (Δ57-486-ATP7B). In contrast to AAV8-ATP7B, AAV8-miniATP7B could be produced at high titers and was able to restore copper homeostasis in 6- and 12-week-old male and female WD mice. In addition, a recently developed synthetic AAV vector, AAVAnc80, carrying the miniATP7B gene was similarly effective at preventing liver damage, restoring copper homeostasis, and improving survival 1 year after treatment. Transduction of approximately 20% of hepatocytes was sufficient to normalize copper homeostasis, suggesting that corrected hepatocytes are acting as a sink to eliminate excess of copper. Importantly, administration of AAVAnc80-miniATP7B was safe in healthy mice and did not result in copper deficiency. Conclusion: In summary, gene therapy using an optimized therapeutic cassette in different AAV systems provides long-term correction of copper metabolism regardless of sex or stage of disease in a clinically relevant WD mouse model. These results pave the way for the implementation of gene therapy in WD patients.
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http://dx.doi.org/10.1002/hep.30535DOI Listing
July 2019

Exosome origin determines cell targeting and the transfer of therapeutic nanoparticles towards target cells.

J Nanobiotechnology 2019 Jan 25;17(1):16. Epub 2019 Jan 25.

Department of Chemical Engineering, Aragon Institute of Nanoscience (INA), University of Zaragoza, Campus Río Ebro-Edificio I+D, C/ Mariano Esquillor S/N, 50018, Zaragoza, Spain.

Background: Exosomes are considered key elements for communication between cells, but very little is known about the mechanisms and selectivity of the transference processes involving exosomes released from different cells.

Results: In this study we have investigated the transfer of hollow gold nanoparticles (HGNs) between different cells when these HGNs were loaded within exosomes secreted by human placental mesenchymal stem cells (MSCs). These HGNs were successfully incorporated in the MSCs exosome biogenesis pathway and released as HGNs-loaded exosomes. Time-lapse microscopy and atomic emission spectroscopy allowed us to demonstrate the selective transfer of the secreted exosomes only to the cell type of origin when studying different cell types including cancer, metastatic, stem or immunological cells.

Conclusions: In this study we demonstrate the selectivity of in vitro exosomal transfer between certain cell types and how this phenomenon can be exploited to develop new specific vectors for advanced therapies. Specifically, we show how this preferential uptake can be leveraged to selectively induce cell death by light-induced hyperthermia only in cells of the same type as those producing the corresponding loaded exosomes. We describe how the exosomes are preferentially transferred to some cell types but not to others, thus providing a better understanding to design selective therapies for different diseases.
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http://dx.doi.org/10.1186/s12951-018-0437-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6346572PMC
January 2019

The in vivo effects of silver nanoparticles on terrestrial isopods, Porcellio scaber, depend on a dynamic interplay between shape, size and nanoparticle dissolution properties.

Analyst 2019 Jan;144(2):488-497

Department of Biology, Biotechnical Faculty, University of Ljubljana, Ljubljana, Slovenia.

The present work aims to study the effects that acute exposure to low concentrations of silver nanoparticles (AgNPs) cause in digestive glands of terrestrial isopods (Porcellio scaber). The experiments were designed to integrate different analytical techniques, such as transmission electron microscopy, atomic absorption spectroscopy, proton induced X-ray emission, and Fourier transform IR imaging (FTIRI), in order to gain a comprehensive insight into the process from the AgNPs' synthesis to their interaction with biological tissues in vivo. To this aim, terrestrial isopods were fed with AgNPs having different shapes, sizes, and concentrations. For all the tested conditions, no toxicity at the whole organism level was observed after 14 days of exposure. However, FTIRI showed that AgNPs caused detectable local changes in proteins, lipids, nucleic acids and carbohydrates at the tissue level, to an extent dependent on the interplay of the AgNPs' properties: shape, size, concentration and dissolution of ions from them.
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http://dx.doi.org/10.1039/c8an01387jDOI Listing
January 2019

Cleavable and thermo-responsive hybrid nanoparticles for on-demand drug delivery.

J Colloid Interface Sci 2019 Jan 23;533:171-181. Epub 2018 Aug 23.

Department of Chemical Engineering, Aragon Institute of Nanoscience (INA), University of Zaragoza, Campus Río Ebro-Edificio I+D, C/ Poeta Mariano Esquillor S/N, 50018 Zaragoza, Spain; Aragon Health Research Institute (IIS Aragón), 50009 Zaragoza, Spain; Networking Research Center on Bioengineering, Biomaterials and Nanomedicine, CIBER-BBN, 28029 Madrid, Spain.

By combining the photothermal ability of copper sulphide nanoparticles (NPs) upon excitation with Near Infrared (NIR) Light and the thermo-responsive properties of the homemade oligo (ethylene glycol) methyl ether methacrylate copolymer we have obtained fragmentable nanocomposites able to release a carried drug on-demand after NIR-light triggering. A complete physico-chemical characterization of the resulting nanoparticles has been carried out and their degradation assessed at different temperatures. Herein, we have also evaluated the drug loading capacity of those nanoparticles and the temperature dependence in their drug release kinetics using bupivacaine hydrochloride as a model drug. For those hybrid nanoparticles, subcytotoxic doses on four different cell lines and their potential interference in cell metabolism, induction of apoptosis, and cell cycle have been evaluated by Alamar Blue fluorometry and flow cytometry.
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http://dx.doi.org/10.1016/j.jcis.2018.08.069DOI Listing
January 2019

Pro-angiogenic near infrared-responsive hydrogels for deliberate transgene expression.

Acta Biomater 2018 09 9;78:123-136. Epub 2018 Aug 9.

University Hospital La Paz-IdiPAZ, Paseo de la Castellana 261, Madrid 28046, Spain; CIBER de Bioingeniería, Biomateriales y Nanomedicina, CIBER-BBN, Spain. Electronic address:

CuS nanoparticles (CuSNP) are degradable, readily prepared, inexpensive to produce and efficiently cleared from the body. In this work, we explored the feasibility of CuSNP to function as degradable near infrared (NIR) nanotransducers within fibrin-based cellular scaffolds. To prepare NIR-responsive CuSNP hydrogels, fibrinogen was dissolved in cell culture medium and supplemented with aqueous dispersions of CuSNP. Fibrinogen polymerization was catalyzed by the addition of thrombin. In some experiments, HUVEC, C3H/10T1/2 or C3H/10T1/2-fLuc cells, that harbor a heat-activated and rapamycin-dependent gene switch for regulating the expression of firefly luciferase transgene, were incorporated to the sol phase of the hydrogel. For in vivo experiments, hydrogels were injected subcutaneously in the back of adult C3H/HeN mice. Upon NIR irradiation, CuSNP hydrogels allowed heat-inducible and rapamycin-dependent transgene expression in cells contained therein, in vitro and in vivo. C3H/10T1/2 cells cultured in CuSNP hydrogels increased metabolic activity, survival rate and fibrinolytic activity, which correlated with changes at the transcriptome level. Media conditioned by CuSNP hydrogels increased viability of HUVEC which formed pseudocapillary structures and remodeled protein matrix when entrapped within these hydrogels. After long-term implantation, the skin patches that covered the CuSNP hydrogels showed increased capillary density which was not detected in mice implanted with matrices lacking CuSNP. In summary, NIR-responsive scaffolds harboring CuSNP offer compelling features in the tissue engineering field, as degradable implants with enhanced integration capacity in host tissues that can provide remote controlled deployment of therapeutic gene products.

Statement Of Significance: Hydrogels composed of fibrin embedding copper sulfide nanoparticles (CuSNP) efficiently convert incident near infrared (NIR) energy into heat and can function as cellular scaffolding. NIR laser irradiation of CuSNP hydrogels can be employed to remotely induce spatiotemporal patterns of transgene expression in genetically engineered multipotent stem cells. CuSNP incorporation in hydrogel architecture accelerates the cell-mediated degradation of the fibrin matrix and induces pro-angiogenic responses that may facilitate the integration of these NIR-responsive scaffolds in host tissues. CuSNP hydrogels that harbor cells capable of controlled expression of therapeutic gene products may be well suited for tissue engineering as they are biodegradable, enhance implant vascularization and can be used to deploy growth factors in a desired spatiotemporal fashion.
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http://dx.doi.org/10.1016/j.actbio.2018.08.006DOI Listing
September 2018