Person: Junquera González, María Elena
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First Name
María Elena
Last Name
Junquera González
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Ciencias Químicas
Department
Química Física
Area
Química Física
Identifiers
16 results
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Now showing 1 - 10 of 16
Item Supramolecular Control over the Interparticle Distance in Gold Nanoparticle Arrays by Cyclodextrin Polyrotaxanes(Nanomaterials, 2018) Coelho, Joao Paulo; Osío Barcina, José De Jesús; Junquera González, María Elena; Aicart Sospedra, Emilio; Tardajos Rodríguez, Gloria; Gómez Graña, Sergio; Cruz Gil, Pablo; Salgado, Cástor; Díaz Núñez, Pablo; Peña Rodríguez, Ovidio; Guerrero Martínez, AndrésAmphiphilic nonionic ligands, synthesized with a fixed hydrophobic moiety formed by a thiolated alkyl chain and an aromatic ring, and with a hydrophilic tail composed of a variable number of oxyethylene units, were used to functionalize spherical gold nanoparticles (AuNPs) in water. Steady-state and time-resolved fluorescence measurements of the AuNPs in the presence of α-cyclodextrin (α-CD) revealed the formation of supramolecular complexes between the ligand and macrocycle at the surface of the nanocrystals. The addition of α-CD induced the formation of inclusion complexes with a high apparent binding constant that decreased with the increasing oxyethylene chain length. The formation of polyrotaxanes at the surface of AuNPs, in which many α-CDs are trapped as hosts on the long and linear ligands, was demonstrated by the formation of large and homogeneous arrays of self-assembled AuNPs with hexagonal close packing, where the interparticle distance increased with the length of the oxyethylene chain. The estimated number of α-CDs per polyrotaxane suggests a high rigidization of the ligand upon complexation, allowing for nearly perfect control of the interparticle distance in the arrays. This degree of supramolecular control was extended to arrays formed by AuNPs stabilized with polyethylene glycol and even to binary arrays. Electromagnetic simulations showed that the enhancement and distribution of the electric field can be finely controlled in these plasmonic arrays.Item A Non-Viral Plasmid DNA Delivery System Consisting on a Lysine-Derived Cationic Lipid Mixed with a Fusogenic Lipid(Pharmaceutics, 2019) Martínez Negro, María; Sánchez Arribas, Natalia; Guerrero Martínez, Andrés; Moyá, María Luisa; Tros de Ilarduya, Conchita; Mendicuti, Francisco; Aicart Sospedra, Emilio; Junquera González, María ElenaThe insertion of biocompatible amino acid moieties in non-viral gene nanocarriers is an attractive approach that has been recently gaining interest. In this work, a cationic lipid, consisting of a lysine-derived moiety linked to a C12 chain (LYCl) was combined with a common fusogenic helper lipid (DOPE) and evaluated as a potential vehicle to transfect two plasmid DNAs (encoding green fluorescent protein GFP and luciferase) into COS-7 cells. A multidisciplinary approach has been followed: (i) biophysical characterization based on zeta potential, gel electrophoresis, small-angle X-ray scattering (SAXS), and cryo-transmission electronic microscopy (cryo-TEM); (ii) biological studies by fluorescence assisted cell sorting (FACS), luminometry, and cytotoxicity experiments; and (iii) a computational study of the formation of lipid bilayers and their subsequent stabilization with DNA. The results indicate that LYCl/DOPE nanocarriers are capable of compacting the pDNAs and protecting them efficiently against DNase I degradation, by forming Lα lyotropic liquid crystal phases, with an average size of ~200 nm and low polydispersity that facilitate the cellular uptake process. The computational results confirmed that the LYCl/DOPE lipid bilayers are stable and also capable of stabilizing DNA fragments via lipoplex formation, with dimensions consistent with experimental values. The optimum formulations (found at 20% of LYCl content) were able to complete the transfection process efficiently and with high cell viabilities, even improving the outcomes of the positive control Lipo2000*.Item Why is less cationic lipid required to prepare lipoplexes from plasmid DNA than linear DNA in gene therapy?(Journal of the American Chemical Society, 2011) Muñoz Úbeda, Mónica; Misra, Santosh ; Barrán-Berdón, Ana; Aicart-Ramos, Clara; Sierra, María ; Biswas, Joydeep; Kondaiah, Paturu; Junquera González, María Elena; Bhattacharya, Santanu; Aicart Sospedra, EmilioThe most important objective of the present study was to explain why cationic lipid (CL)-mediated delivery of plasmid DNA (pDNA) is better than that of linear DNA in gene therapy, a question that, until now, has remained unanswered. Herein for the first time we experimentally show that for different types of CLs, pDNA, in contrast to linear DNA, is compacted with a large amount of its counterions, yielding a lower effective negative charge. This feature has been confirmed through a number of physicochemical and biochemical investigations. This is significant for both in vitro and in vivo transfection studies. For an effective DNA transfection, the lower the amount of the CL, the lower is the cytotoxicity. The study also points out that it is absolutely necessary to consider both effective charge ratios between CL and pDNA and effective pDNA charges, which can be determined from physicochemical experiments.Item Gemini-Based Lipoplexes Complement the Mitochondrial Phenotype in MFN1-Knockout Mouse Embryonic Fibroblasts(Molecular Pharmaceutics, 2019) Muñoz Úbeda, Mónica; Tolosa Díaz, Andrés; Junquera González, María Elena; Bhattacharya, Santanu; Aicart Sospedra, Emilio; Natale, Paolo; López-Montero, IvánMitochondria form a dynamic network of constantly dividing and fusing organelles. The balance between these antagonistic processes is crucial for normal cellular function and requires the action of specialized proteins. The mitochondrial membrane proteins mitofusin 1 (Mfn1) and mitofusin 2 (Mfn2) are responsible for the fusion of the outer membrane of adjacent mitochondria. Mutations within Mfn1 or Mfn2 impair mitochondrial fusion and lead to some severe mitochondrial dysfunctions and mitochondrial diseases (MDs). A characteristic phenotype of cells carrying defective Mfn1 or Mfn2 is the presence of a highly fragmented mitochondrial network. Here, we use a biocompatible mixture of lipids, consisting on synthetic gemini cationic lipids (GCLs) and the zwitterionic phospholipid (DOPE), to complex, transport, and deliver intact copies of MFN1 gene into MFN1-Knockout mouse embryonic fibroblasts (MFN1-KO MEFs). We demonstrate that the GCL/DOPE-DNA lipoplexes are able to introduce the intact MFN1 gene into the cells and ectopically produce functional Mfn1. A four-fold increase of the Mfn1 levels is necessary to revert the MFN1-KO phenotype and to partially restore a mitochondrial network. This phenotype complementation was correlated with the transfection of GCL/ DOPE-MFN1 lipoplexes that exhibited a high proportion of highly packaged hexagonal phase. GCL/DOPE-DNA lipoplexes are formulated as efficient therapeutic agents against MDs.Item How does the spacer length of cationic gemini lipids influence thelLipoplex formation with plasmid DNA? Physicochemical and biochemical characterizations and their relevance in gene therapy(Biomacromolecules, 2012) Muñoz Úbeda, Mónica; Misra, Santosh K.; Barran-Berdón, Ana L; Datta, Sougata; Aicart-Ramos, Clara; Castro-Hartmann, Pablo; Kondaiah, Paturu; Junquera González, María Elena; Bhattacharya, Santanu; Aicart Sospedra, EmilioLipoplexes formed by the pEGFP-C3 plasmid DNA (pDNA) and lipid mixtures containing cationic gemini surfactant of the 1,2-bis(hexadecyl dimethyl ammonium) alkanes family referred to as C16CnC16, where n = 2, 3, 5, or 12, and the zwitterionic helper lipid, 1,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine (DOPE) have been studied from a wide variety of physical, chemical, and biological standpoints. The study has been carried out using several experimental methods, such as zeta potential, gel electrophoresis, small angle X-ray scattering (SAXS), cryo-TEM, gene transfection, cell viability/cytotoxicity, and confocal fluorescence microscopy. As reported recently in a communication (J. Am. Chem. Soc. 2011, 133, 18014), the detailed physicochemical and biological studies confirm that, in the presence of the studied series lipid mixtures, plasmid DNA is compacted with a large number of its associated Na+ counterions. This in turn yields a much lower effective negative charge, qpDNA − , a value that has been experimentally obtained for each mixed lipid mixture. Consequently, the cationic lipid (CL) complexes prepared with pDNA and CL/DOPE mixtures to be used in gene transfection require significantly less amount of CL than the one estimated assuming a value of qDNA − = −2. This drives to a considerably lower cytotoxicity of the gene vector. Depending on the CL molar composition, α, of the lipid mixture, and the effective charge ratio of the lipoplex, ρeff, the reported SAXS data indicate the presence of two or three structures in the same lipoplex, one in the DOPE-rich region, other in the CL-rich region, and another one present at any CL composition. Cryo-TEM and SAXS studies with C16CnC16/DOPE-pDNA lipoplexes indicate that pDNA is localized between the mixed lipidm bilayers of lamellar structures within a monolayer of ∼2 nm. This is consistent with a highly compacted supercoiled pDNA conformation compared with that of linear DNA. Transfection studies were carried out with HEK293T, HeLa, CHO, U343, and H460 cells. The α and ρeff values for each lipid mixture were optimized on HEK293T cells for transfection, and using these values, the remaining cells were also transfected in absence (-FBS-FBS) and presence (-FBS+FBS) of serum. The transfection efficiency was higher with the CLs of shorter gemini spacers (n = 2 or 3). Each formulation expressed GFP on pDNA transfection and confocal fluorescence microscopy corroborated the results. C16C2C16/DOPE mixtures were the most efficient toward transfection among all the lipid mixtures and, in presence of serum, even better than the Lipofectamine2000, a commercial transfecting agent. Each lipid combination was safe and did not show any significant levels of toxicity. Probably, the presence of two coexisting lamellar structures in lipoplexes synergizes the transfection efficiency of the lipid mixtures which are plentiful in the lipoplexes formed by CLs with short spacer (n = 2, 3) thanm, those with the long spacer (n = 5, 12).Item Experimental and theoretical approach to the sodium decanoate-dodecanoate mixed surfactant system in aqueous solution(Langmuir, 2010) Rodríguez-Pulido, Alberto; Casado, Aitor; Muñoz Úbeda, Mónica; Junquera González, María Elena; Aicart Sospedra, EmilioThe mixed system consisting of two anionic surfactants of identical headgroups but with 10 and 12 carbon atoms on the hydrophobic tail, sodium decanoate (C10Na) and sodium dodecanoate (C12Na), has been studied in aqueous solution at 298.15 K by means of conductivity and fluorescence spectroscopy experiments and from a theoretical point of view. The monomeric and micellar phases of the mixed aggregates were analyzed through the experimental determination of the total critical micelle concentration, cmc*, the degree of ionization of the mixed micelle, β, and the total aggregation number, N*. Results indicate that, compared to the ideal behavior, the mixed system with two anionic surfactants differing only in two methylenes in the hydrophobic tail shows a negative deviation in the cmc* and a positive one in N*. Pure surfactants (C10Na and C12Na) form spherical micelles, but mixed micelles must aggregate with a rodlike shape to allow more surfactant molecules than expected. In addition, rodlike micelles result in more compacted aggregation (i.e., less area per polar head). From the experimental data in this work, several theoretical models for mixed surfactant systems have been checked: Rubingh’s model predicts lower deviations from ideality than Motomura’s model. The stability of the micelles has been analyzed by computing the standard Gibbs energy of micelle formation, ΔGmic,0, of pure and mixed micelles. Results of this work reinforce the feature that mixed systems formed by alkylsurfactants with the same polar head that differ in the hydrocarbon length, usually admitted as roughly ideal systems, may show nonideal behavior. This deviation, being mostly related to the difference in the chain length, Δnc, between surfactants can be analyzed only when very accurate experimental techniques as well as adequate theoretical models are used.Item Effects of a delocalizable cation on the headgroup of gemini lipids on the lipoplex-type nanoaggregates directly formed from plasmid DNA(Biomacromolecules, 2013) Misra, Santosh ; Muñoz Úbeda, Mónica; Datta, Sougata; Barrán-Berdón, Ana L.; Aicart-Ramos, Clara; Castro-Hartmann, Pablo; Kondaiah, Paturu; Junquera González, María Elena; Battacharya, Santanu; Aicart Sospedra, EmilioLipoplex-type nanoaggregates prepared from pEGFP-C3 plasmid DNA (pDNA) and mixed liposomes, with a gemini cationic lipid (CL) [1,2-bis(hexadecyl imidazolium) alkanes], referred as (C16Im)2Cn (where Cn is the alkane spacer length, n = 2, 3, 5, or 12, between the imidazolium heads) and DOPE zwitterionic lipid, have been analyzed by zeta potential, gel electrophoresis, SAXS, cryo-TEM, fluorescence anisotropy, transfection efficiency, fluorescence confocal microscopy, and cell viability/cytotoxicity experiments to establish a structure−biological activity relationship. The study, carried out at several mixed liposome compositions, α, and effective charge ratios, ρeff, of the lipoplex, demonstrates that the transfection of pDNA using CLs initially requires the determination of the effective charge of both. The electrochemical study confirms that CLs with a delocalizable positive charge in their headgroups yield an effective positive charge that is 90% of their expected nominal one, while pDNA is compacted yielding an effective negative charge which is only 10−25% than that of the linear DNA. SAXS diffractograms show that lipoplexes formed by CLs with shorter spacer (n = 2, 3, or 5) present three lamellar structures, two of them in coexistence, while those formed by CL with longest spacer (n = 12) present two additional inverted hexagonal structures. Cryo-TEM micrographs show nanoaggregates with two multilamellar structures, a cluster-type (at low α value) and a fingerprinttype, that coexist with the cluster-type at moderate α composition. The optimized transfection efficiency (TE) of pDNA, in HEK293T, HeLa, and H1299 cells was higher using lipoplexes containing gemini CLs with shorter spacers at low α value. Each lipid formulation did not show any significant levels of toxicity, the reported lipoplexes being adequate DNA vectors for gene therapy and considerably better than both Lipofectamine 2000 and CLs of the 1,2-bis(hexadecyl ammnoniun) alkane series, recently reported.Item Complejos de inclusión de ciclodextrinas en medios micelares(2002) Junquera González, María Elena; Tardajos Rodríguez, Gloria María; Aicart Sospedra, EmilioEn este trabajo se ha llevado a cabo un amplio estudio de los complejos de inclusión que forman las ciclodextrinas con varios surfactantes por la microencapsulacion de los mismos en la cavidad de las primeras. Se ha trabajado con dos ciclodextrinas, la beta-cd y una sustituida de la misma, la 2,6-o-dimetil- -cd, mientras que como surfactantes se han utilizado los bromuro de deciltrimetilamonio (dtab) y hexadeciltrimetilamonio (ctab), el dodecilsulfato de sodio (sds) y el perfluoroctanoato de sodio (spfo). De esta forma, se ha analizado la influencia de la ciclodextrina, la longitud de la cadena de surfactante, la carga del mismo (cationicos o anionicos) y la fluoracion de la cadena en el proceso de complejacion. Para ello, se han utilizado dos técnicas ultrasónicas, la velocidad del sonido y la absorción ultrasónica, además de una técnica conductimetrica y una de fluorescencia inducida por láser. Con toda la información experimental, se ha realizado un estudio termodinamico y cinético del proceso de inclusión y su influencia en el de micelizacion.Item Intercellular Trafficking of Gold Nanostars in Uveal Melanoma Cells for Plasmonic Photothermal Therapy(Nanomaterials, 2020) Ahijado Guzmán, Rubén; Sánchez Arribas, Natalia; Martínez Negro, María; González Rubio, Guillermo; Santiago-Varela, María; Pardo, María; Piñeiro, Antonio; López-Montero, Iván; Junquera González, María Elena; Guerrero Martínez, AndrésEfficient plasmonic photothermal therapies (PPTTs) using non-harmful pulse laser irradiation at the near-infrared (NIR) are a highly sought goal in nanomedicine. These therapies rely on the use of plasmonic nanostructures to kill cancer cells while minimizing the applied laser power density. Cancer cells have an unsettled capacity to uptake, retain, release, and re-uptake gold nanoparticles, thus offering enormous versatility for research. In this work, we have studied such cell capabilities for nanoparticle trafficking and its impact on the effect of photothermal treatments. As our model system, we chose uveal (eye) melanoma cells, since laser-assisted eye surgery is routinely used to treat glaucoma and cataracts, or vision correction in refractive surgery. As nanostructure, we selected gold nanostars (Au NSs) due to their high photothermal efficiency at the near-infrared (NIR) region of the electromagnetic spectrum. We first investigated the photothermal effect on the basis of the dilution of Au NSs induced by cell division. Using this approach, we obtained high PPTT efficiency after several cell division cycles at an initial low Au NS concentration (pM regime). Subsequently, we evaluated the photothermal effect on account of cell division upon mixing Au NS-loaded and non-loaded cells. Upon such mixing, we observed trafficking of Au NSs between loaded and non-loaded cells, thus achieving effective PPTT after several division cycles under low irradiation conditions (below the maximum permissible exposure threshold of skin). Our study reveals the ability of uveal melanoma cells to release and re-uptake Au NSs that maintain their plasmonic photothermal properties throughout several cell division cycles and re-uptake. This approach may be readily extrapolated to real tissue and even to treat in situ the eye tumor itself. We believe that our method can potentially be used as co-therapy to disperse plasmonic gold nanostructures across affected tissues, thus increasing the effectiveness of classic PPTT.Item Gemini Cationic Lipid-Type Nanovectors Suitable for the Transfection of Therapeutic Plasmid DNA Encoding for Pro-Inflammatory Cytokine Interleukin-12(Pharmaceutics, 2021) Sánchez Arribas, Natalia; Martínez Negro, María; Aicart Ramos, Clara; Tros de Ilarduya, Conchita; Aicart Sospedra, Emilio; Guerrero Martínez, Andrés; Junquera González, María ElenaAmple evidence exists on the role of interleukin-12 (IL-12) in the response against many pathogens, as well as on its remarkable antitumor properties. However, the unexpected toxicity and disappointing results in some clinical trials are prompting the design of new strategies and/or vectors for IL-12 delivery. This study was conceived to further endorse the use of gemini cationic lipids (GCLs) in combination with zwitterionic helper lipid DOPE (1,2-dioleoyl-sn-glycero-3-phosphatidyl ethanol amine) as nanovectors for the insertion of plasmid DNA encoding for IL-12 (pCMV-IL12) into cells. Optimal GCL formulations previously reported by us were selected for IL-12-based biophysical experiments. In vitro studies demonstrated efficient pCMV-IL12 transfection by GCLs with comparable or superior cytokine levels than those obtained with commercial control Lipofectamine2000*. Furthermore, the nanovectors did not present significant toxicity, showing high cell viability values. The proteins adsorbed on the nanovector surface were found to be mostly lipoproteins and serum albumin, which are both beneficial to increase the blood circulation time. These outstanding results are accompanied by an initial physicochemical characterization to confirm DNA compaction and protection by the lipid mixture. Although further studies would be necessary, the present GCLs exhibit promising characteristics as candidates for pCMV-IL12 transfection in future in vivo applications.