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Gene therapeutics are promising for treating diseases at the genetic level, with some already validated for clinical use. Recently, nanostructures have emerged for the targeted delivery of genetic material. Nanomaterials, exhibiting advantageous properties such as a high surface-to-volume ratio, biocompatibility, facile functionalization, substantial loading capacity, and tunable physicochemical characteristics, are recognized as non-viral vectors in gene therapy applications. Despite progress, current non-viral vectors exhibit notably low gene delivery efficiency. Progress in nanotechnology is essential to overcome extracellular and intracellular barriers in gene delivery. Specific nanostructures such as carbon nanotubes (CNTs), carbon quantum dots (CQDs), nanodiamonds (NDs), and similar carbon-based structures can accommodate diverse genetic materials such as plasmid DNA (pDNA), messenger RNA (mRNA), small interference RNA (siRNA), micro RNA (miRNA), and antisense oligonucleotides (AONs). To address challenges such as high toxicity and low transfection efficiency, advancements in the features of carbon-based nanostructures (CBNs) are imperative. This overview delves into three types of CBNs employed as vectors in drug/gene delivery systems, encompassing their synthesis methods, properties, and biomedical applications. Ultimately, we present insights into the opportunities and challenges within the captivating realm of gene delivery using CBNs.

Background: Lenvatinib, a tyrosine kinase inhibitor (TKI) approved for the treatment of progressive and radioactive iodine (RAI)-refractory differentiated thyroid cancer (DTC), is associated with significant adverse effects that can be partially mitigated through the development of novel drug formulations. The utilization of nanoparticles presents a viable option, as it allows for targeted drug delivery, reducing certain side effects and enhancing the overall quality of life for patients. This study aimed to produce and assess, both in vitro and in vivo, the cytotoxicity, biodistribution, and therapeutic efficacy of lenvatinib-loaded PLGA nanoparticles (NPs), both with and without decoration using antibody conjugation (cetuximab), as a novel therapeutic approach for managing aggressive thyroid tumors. Methods: Poly(lactic-co-glycolic acid) nanoparticles (NPs), decorated with or without anti-EGFR, were employed as a lenvatinib delivery system. These NPs were characterized for size distribution, surface morphology, surface charge, and drug encapsulation efficiency. Cytotoxicity was evaluated through MTT assays using two cellular models, one representing normal thyroid cells (Nthy-ori 3-1) and the other representing anaplastic thyroid cells (CAL-62). Additionally, an in vivo xenograft mouse model was established to investigate biodistribution and therapeutic efficacy following intragastric administration. Results: The NPs demonstrated success in terms of particle size, polydispersity index (PDI), zeta potential, morphology, encapsulation efficiency, and cetuximab distribution across the surface. In vitro analysis revealed cytotoxicity in both cellular models with both formulations, but only the decorated NPs achieved an ID50 value in CAL-62 cells. Biodistribution analysis following intragastric administration in xenografted thyroid mice demonstrated good stability in terms of intestinal barrier function and tumor accumulation. Both formulations were generally well tole

Gene therapy and optogenetics are becoming promising tools for treating several nervous system pathologies. Currently, most of these approaches use viral vectors to transport the genetic material inside the cells, but viruses present some potential risks, such as marked immunogenicity, insertional mutagenesis, and limited insert gene size. In this framework, non-viral nanoparticles, such as niosomes, are emerging as possible alternative tools to deliver genetic material, avoiding the aforementioned problems. To determine their suitability as vectors for optogenetic therapies in this work, we tested three different niosome formulations combined with three optogenetic plasmids in rat cortical neurons in vitro. All niosomes tested successfully expressed optogenetic channels, which were dependent on the ratio of niosome to plasmid, with higher concentrations yielding higher expression rates. However, we found changes in the dendritic morphology and electrophysiological properties of transfected cells, especially when we used higher concentrations of niosomes. Our results highlight the potential use of niosomes for optogenetic applications and suggest that special care must be taken to achieve an optimal balance of niosomes and nucleic acids to achieve the therapeutic effects envisioned by these technologies.

227 p., The integration of novel compounds into formulations may open up new avenues and opportunities for improving the treatment of various diseases using a genetherapytechnique. To this purpose, my thesis aimed to study the impact of incorporation of biomaterials such sphingolipids or nanodiamonds as a helper component into a cationic niosome formulation for gene delivery, the formulations and related complexes were studied in terms of particle size,surface charge, dispersity, and morphology. Furthermore, biological research were conducted inboth in vitro and in vivo experimental trials to evaluate cellular uptake, intracellular trafficking pathways, and transfection efficiency.

Nanodiamonds were combined with niosome, and resulting formulations were named as nanodiasomes, which were evaluated in terms of physicochemical features, cellular internalization, cell viability and transfection efficiency both in in vitro and in in vivo conditions. Such parameters were analyzed at 4 and 25 °C, and at 15 and 30 days after their elaboration. Nanodiasomes showed a particle size of 128 nm that was maintained over time inside the ± 10% of deviation, unless after 30 days of storage at 25 °C. Something similar occurred with the initial zeta potential value, 35.2 mV, being both formulations more stable at 4 °C. The incorporation of nanodiamonds into niosomes resulted in a 4-fold increase of transfection efficiency that was maintained over time at 4 and 25 °C. In vivo studies reported high transgene expression of nanodiasomes after subretinal and intravitreal administration in mice, when injected freshly prepared and after 30 days of storage at 4 °C.

[EN] Central nervous system (CNS) diseases, especially acute ischemic events and neurodegenerative disorders, constitute a public health problem with no effective treatments to allow a persistent solution. Failed therapies targeting neuronal recovery have revealed the multifactorial and intricate pathophysiology underlying such CNS disorders as ischemic stroke, Alzheimers disease, amyotrophic lateral sclerosis, vascular Parkisonism, vascular dementia, and aging, in which cerebral microvasculature impairment seems to play a key role. In fact, a reduction in vessel density and cerebral blood flow occurs in these scenarios, contributing to neuronal dysfunction and leading to loss of cognitive function. In this review, we provide an overview of healthy brain microvasculature structure and function in health and the effect of the aforementioned cerebral CNS diseases. We discuss the emerging new therapeutic opportunities, and their delivery approaches, aimed at recovering brain vascularization in this context. SIGNIFICANCE STATEMENT: The lack of effective treatments, mainly focused on neuron recovery, has prompted the search of other therapies to treat cerebral central nervous system diseases. The disruption and degeneration of cerebral microvasculature has been evidenced in neurodegenerative diseases, stroke, and aging, constituting a potential target for restoring vascularization, neuronal functioning, and cognitive capacities by the development of therapeutic pro-angiogenic strategies.

[EN] Nanodiamonds (NDs) are promising materials for gene delivery because of their unique physicochemical and biological features, along with their possibility of combination with other nonviral systems. Our aim was to evaluate the biophysical performance of NDs as helper components of niosomes, named nanodiasomes, to address a potential nonviral gene delivery nanoplatform for therapeutic applications in central nervous system (CNS) diseases. Nanodiasomes, niosomes, and their corresponding complexes, obtained after genetic material addition at different ratios (w/w), were evaluated in terms of physicochemical properties, cellular uptake, intracellular disposition, biocompatibility, and transfection efficiency in HEK-293 cells. Nanodiasomes, niosomes, and complexes fulfilled the physicochemical features for gene therapy applications. Biologically, the incorporation of NDs into niosomes enhanced 75% transfection efficiency (p < 0.001) and biocompatibility (p < 0.05) to values over 90%, accompanied by a higher cellular uptake (p < 0.05). Intracellular trafficking analysis showed higher endocytosis via clathrins (p < 0.05) in nanodiaplexes compared with nioplexes, followed by higher lysosomal colocalization (p < 0.05), that coexisted with endosomal escape properties, whereas endocytosis mediated by caveolae was the most efficient pathway in the case of nanodiaplexes. Moreover, studies in CNS primary cells revealed that nanodiaplexes successfully transfected neuronal and retinal cells. This proof-of-concept study points out that ND integration into niosomes represents an encouraging nonviral nanoplatform strategy for the treatment of CNS diseases by gene therapy.

[EN] Nanodiamonds (NDs) are promising materials for gene delivery because of their unique physicochemical and biological features, along with their possibility of combination with other nonviral systems. Our aim was to evaluate the biophysical performance of NDs as helper components of niosomes, named nanodiasomes, to address a potential nonviral gene delivery nanoplatform for therapeutic applications in central nervous system (CNS) diseases. Nanodiasomes, niosomes, and their corresponding complexes, obtained after genetic material addition at different ratios (w/w), were evaluated in terms of physicochemical properties, cellular uptake, intracellular disposition, biocompatibility, and transfection efficiency in HEK-293 cells. Nanodiasomes, niosomes, and complexes fulfilled the physicochemical features for gene therapy applications. Biologically, the incorporation of NDs into niosomes enhanced 75% transfection efficiency (p < 0.001) and biocompatibility (p < 0.05) to values over 90%, accompanied by a higher cellular uptake (p < 0.05). Intracellular trafficking analysis showed higher endocytosis via clathrins (p < 0.05) in nanodiaplexes compared with nioplexes, followed by higher lysosomal colocalization (p < 0.05), that coexisted with endosomal escape properties, whereas endocytosis mediated by caveolae was the most efficient pathway in the case of nanodiaplexes. Moreover, studies in CNS primary cells revealed that nanodiaplexes successfully transfected neuronal and retinal cells. This proof-of-concept study points out that ND integration into niosomes represents an encouraging nonviral nanoplatform strategy for the treatment of CNS diseases by gene therapy.

[EN] Central nervous system (CNS) diseases, especially acute ischemic events and neurodegenerative disorders, constitute a public health problem with no effective treatments to allow a persistent solution. Failed therapies targeting neuronal recovery have revealed the multifactorial and intricate pathophysiology underlying such CNS disorders as ischemic stroke, Alzheimers disease, amyotrophic lateral sclerosis, vascular Parkisonism, vascular dementia, and aging, in which cerebral microvasculature impairment seems to play a key role. In fact, a reduction in vessel density and cerebral blood flow occurs in these scenarios, contributing to neuronal dysfunction and leading to loss of cognitive function. In this review, we provide an overview of healthy brain microvasculature structure and function in health and the effect of the aforementioned cerebral CNS diseases. We discuss the emerging new therapeutic opportunities, and their delivery approaches, aimed at recovering brain vascularization in this context. SIGNIFICANCE STATEMENT: The lack of effective treatments, mainly focused on neuron recovery, has prompted the search of other therapies to treat cerebral central nervous system diseases. The disruption and degeneration of cerebral microvasculature has been evidenced in neurodegenerative diseases, stroke, and aging, constituting a potential target for restoring vascularization, neuronal functioning, and cognitive capacities by the development of therapeutic pro-angiogenic strategies.

Retinitis pigmentosa (RP) is the most common inherited retinal dystrophy causing progressive vision loss. It is accompanied by chronic and sustained inflammation, including M1 microglia activation. This study evaluated the effect of an essential fatty acid (EFA) supplement containing specialized pro-resolving mediators (SPMs), on retinal degeneration and microglia activation in rd10 mice, a model of RP, as well as on LPS-stimulated BV2 cells. The EFA supplement was orally administered to mice from postnatal day (P)9 to P18. At P18, the electrical activity of the retina was examined by electroretinography (ERG) and innate behavior in response to light were measured. Retinal degeneration was studied via histology including the TUNEL assay and microglia immunolabeling. Microglia polarization (M1/M2) was assessed by flow cytometry, qPCR, ELISA and histology. Redox status was analyzed by measuring antioxidant enzymes and markers of oxidative damage. Interestingly, the EFA supplement ameliorated retinal dysfunction and degeneration by improving ERG recording and sensitivity to light, and reducing photoreceptor cell loss. The EFA supplement reduced inflammation and microglia activation attenuating M1 markers as well as inducing a shift to the M2 phenotype in rd10 mouse retinas and LPS-stimulated BV2 cells. It also reduced oxidative stress markers of lipid peroxidation and carbonylation. These findings could open up new therapeutic opportunities based on resolving inflammation with oral supplementation with SPMs such as the EFA supplement.

[EN] Nanodiamonds (NDs) are promising materials for gene delivery because of their unique physicochemical and biological features, along with their possibility of combination with other nonviral systems. Our aim was to evaluate the biophysical performance of NDs as helper components of niosomes, named nanodiasomes, to address a potential nonviral gene delivery nanoplatform for therapeutic applications in central nervous system (CNS) diseases. Nanodiasomes, niosomes, and their corresponding complexes, obtained after genetic material addition at different ratios (w/w), were evaluated in terms of physicochemical properties, cellular uptake, intracellular disposition, biocompatibility, and transfection efficiency in HEK-293 cells. Nanodiasomes, niosomes, and complexes fulfilled the physicochemical features for gene therapy applications. Biologically, the incorporation of NDs into niosomes enhanced 75% transfection efficiency (p < 0.001) and biocompatibility (p < 0.05) to values over 90%, accompanied by a higher cellular uptake (p < 0.05). Intracellular trafficking analysis showed higher endocytosis via clathrins (p < 0.05) in nanodiaplexes compared with nioplexes, followed by higher lysosomal colocalization (p < 0.05), that coexisted with endosomal escape properties, whereas endocytosis mediated by caveolae was the most efficient pathway in the case of nanodiaplexes. Moreover, studies in CNS primary cells revealed that nanodiaplexes successfully transfected neuronal and retinal cells. This proof-of-concept study points out that ND integration into niosomes represents an encouraging nonviral nanoplatform strategy for the treatment of CNS diseases by gene therapy.

[EN] Central nervous system (CNS) diseases, especially acute ischemic events and neurodegenerative disorders, constitute a public health problem with no effective treatments to allow a persistent solution. Failed therapies targeting neuronal recovery have revealed the multifactorial and intricate pathophysiology underlying such CNS disorders as ischemic stroke, Alzheimers disease, amyotrophic lateral sclerosis, vascular Parkisonism, vascular dementia, and aging, in which cerebral microvasculature impairment seems to play a key role. In fact, a reduction in vessel density and cerebral blood flow occurs in these scenarios, contributing to neuronal dysfunction and leading to loss of cognitive function. In this review, we provide an overview of healthy brain microvasculature structure and function in health and the effect of the aforementioned cerebral CNS diseases. We discuss the emerging new therapeutic opportunities, and their delivery approaches, aimed at recovering brain vascularization in this context. SIGNIFICANCE STATEMENT: The lack of effective treatments, mainly focused on neuron recovery, has prompted the search of other therapies to treat cerebral central nervous system diseases. The disruption and degeneration of cerebral microvasculature has been evidenced in neurodegenerative diseases, stroke, and aging, constituting a potential target for restoring vascularization, neuronal functioning, and cognitive capacities by the development of therapeutic pro-angiogenic strategies.

In the present work, single wall carbon nanotubes (SWCNT) were successively functionalized with phospholipid DSPE-PEG carboxylic acid, and then, with ethylenediamine (EDA), to obtain double functionalized single wall carbon nanotube (DFSWCNT). Then, DFSWCNT was applied as a carrier for delivering amphotericin B (Amb) and EGFP plasmid. FSWCNT’s concentration obtained via UV–visible analysis was 0.99 mg/mL. The TGA analysis results provided the lost weights of DSPE-PEG-COOH, EDA, Amb and SWCNT impurities. XPS results showed that carbon atoms’ percentage decreased during the functionalization processes from 97.2% (SWCNT) to 76.4% (FSWCNT) and 69.9% (DFSWNCT). Additionally, the oxygen atoms’ percentage increased from 2.3% (SWCNT) to 21% and 22.5% for FSWCNT and DFSWCNT, respectively. New bonds such as C–N and N–C=O appeared in the synthesized nanocarrier. The IG/ID ratio in Raman analysis decreased from 7.15 (SWCNT) to 4.08 (FSWCNT). The amount of Amb released to phosphate buffer saline medium was about 33% at pH = 5.5 and 75% at pH = 7.4 after 48 h. CCK8 results confirmed that the toxicity of functionalized SWCNT had decreased. In a 2:1 ratio of DFSWCNT/EGFP plasmid, the cell viability (87%) and live transfected cells (56%) were at their maximum values. The results indicate that carbon nanotubes have the potential to be applied as drug/gene delivery systems with outstanding properties such as high loading capacity and easy penetration to cell membrane.

Lipid nanocarriers, such as niosomes, are considered attractive candidates for non-viral gene delivery due to their suitable biocompatibility and high versatility. In this work, we studied the influence of incorporating chloroquine in niosomes biophysical performance, as well as the effect of non-ionic surfactant composition and protocol of incorporation in their biophysical performance. An exhaustive comparative evaluation of three niosome formulations differing in these parameters was performed, which included the analysis of their thermal stability, rheological behavior, mean particle size, dispersity, zeta potential, morphology, membrane packing capacity, affinity to bind DNA, ability to release and protect the genetic material, buffering capacity and ability to escape from artificially synthesized lysosomes. Finally, in vitro biological studies were, also, performed in order to determine the compatibility of the formulations with biological systems, their transfection efficiency and transgene expression. Results revealed that the incorporation of chloroquine in niosome formulations improved their biophysical properties and the transfection efficiency, while the substitution of one of the non-ionic surfactants and the phase of addition resulted in less biophysical variations. Of note, the present work provides several biophysical parameters and characterization strategies that could be used as gold standard for gene therapy nanosystems evaluation.

[EN]The aim was to evaluate relevant biophysic processes related to the physicochemical features and gene transfection mechanism when sphingolipids are incorporated into a cationic niosome formulation for non-viral gene delivery to central nervous system. For that, two formulations named niosphingosomes and niosomes devoid of sphingolipid extracts, as control, were developed by the oil-in water emulsion technique. Both formulations and the corresponding complexes, obtained upon the addition of the reporter EGFP plasmid, were physicochemically and biologically characterized and evaluated. Compared to niosomes, niosphingosomes, and the corresponding complexes decreased particle size and increased superficial charge. Although there were not significant differences in the cellular uptake, cell viability and transfection efficiency increased when human retinal pigment epithelial (ARPE-19) cells were exposed to niosphingoplexes. Endocytosis via caveolae decreased in the case of niosphingoplexes, which showed higher co-localization with lysosomal compartment, and endosomal escape properties. Moreover, niosphingoplexes transfected not only primary central nervous system cells, but also different cells in mouse retina, depending on the administration route, and brain cortex. These preliminary results suggest that niosphingosomes represent a promising non-viral vector formulation purposed for the treatment of both retinal and brain diseases by gene therapy approach.

The cell-based approach in gene therapy arises as a promising strategy to provide safe, targeted, and efficient gene delivery. Owing to their unique features, as homing and tumor-tropism, mesenchymal stem cells (MSCs) have recently been introduced as an encouraging vehicle in gene therapy. Nevertheless, non-viral transfer of nucleic acids into MSCs remains limited due to various factors related to the main stakeholders of the process (e.g., nucleic acids, carriers, or cells). In this review, we have summarized the main types of nucleic acids used to transfect MSCs, the pros and cons, and applications of each. Then, we have emphasized on the most efficient lipid-based carriers for nucleic acids to MSCs, their main features, and some of their applications. While a myriad of studies have demonstrated the therapeutic potential for engineered MSCs therapy in various illnesses, optimization for clinical use is an ongoing challenge. On the way of improvement, genetically modified MSCs have been combined with various novel techniques and tools (e.g., exosomes, spheroids, 3D-Bioprinting, etc.,) aiming for more efficient and safe applications in biomedicine.

Efficient delivery of genetic material into cells is a critical process to translate gene therapy into clinical practice. In this sense, the increased knowledge acquired during past years in the molecular biology and nanotechnology fields has contributed to the development of different kinds of non-viral vector systems as a promising alternative to virus-based gene delivery counterparts. Consequently, the development of non-viral vectors has gained attention, and nowadays, gene delivery mediated by these systems is considered as the cornerstone of modern gene therapy due to relevant advantages such as low toxicity, poor immunogenicity and high packing capacity. However, despite these relevant advantages, non-viral vectors have been poorly translated into clinical success. This review addresses some critical issues that need to be considered for clinical practice application of non-viral vectors in mainstream medicine, such as efficiency, biocompatibility, long-lasting effect, route of administration, design of experimental condition or commercialization process. In addition, potential strategies for overcoming main hurdles are also addressed. Overall, this review aims to raise awareness among the scientific community and help researchers gain knowledge in the design of safe and efficient non-viral gene delivery systems for clinical applications to progress in the gene therapy field.

26 p. : il. -- Bibliogr.: p. 24-25, Alzheimer gaixotasuna, gaixotasun neurologiko bat da, eta, dementzia da bere sintomarik nagusiena. Adinarekin agertzen den gaixotasun multifaktoriala da. Gaixotasun honen hipotesi ezagunena ß-amiloide metaketa da; baina, azken urteetan beste hainbat hipotesi aurkitzen saiatu izan dira, ß-amiloide plaka horien gain eragiten duten tratamenduen eraginaren ondorioz ikusi delako ez dela beti gaixotasunaren hobekuntza ikusten. Beraz, beste hainbat hipotesi aztertu dira, adibidez, garuneko baskularizazioaren gutxipena. Azken faktore hau azken urteetan garrantzia hartzen ari da, eta tratamendua faktore honetan oinarrituz garatzea da helburua. Gaur egun, merkatuan dauden tratamenduak sintomen gain soilik eragiten baitute eta ez gaixotasunaren garapenean. VEGF faktoreak eragin neurobabesle eta angiogenikoa dituenez Alzheimerren gaixotasunean eraginkorra izan daitekeela uste da. Horregatik, maila preklinikoan ikerketa ezberdinak egin ondoren eta horiek adierazitako emaitza positiboak aztertu ondoren, etorkizunean eraginkorra izan daitekeela uste da. Eman-bidea ere ikertzeko beste atal garrantzitsu bat izan da gaixotasun honetan, gaixotasun neurologiko bat denez ez baita horren erraza farmakoak bertara helaraztea. Kasu honetan, sudur-bidea aztertu da hesi hematoentzefalikoa gainditzeko gai izan daitekeen eman-bide berritzaile eta ez-inbaditzaile gisa, etorkizunerako aukera bat izan daitekeelakoan.

Three-dimensional (3D) printing is a game changer technology that holds great promise for a wide variety of biomedical applications, including ophthalmology. Through this emerging technique, specific eye tissues can be custom-fabricated in a flexible and automated way, incorporating different cell types and biomaterials in precise anatomical 3D geometries. However, and despite the great progress and possibilities generated in recent years, there are still challenges to overcome that jeopardize its clinical application in regular practice. The main goal of this review is to provide an in-depth understanding of the current status and implementation of 3D bioprinting technology in the ophthalmology field in order to manufacture relevant tissues such as cornea, retina and conjunctiva. Special attention is paid to the description of the most commonly employed bioprinting methods, and the most relevant eye tissue engineering studies performed by 3D bioprinting technology at preclinical level. In addition, other relevant issues related to use of 3D bioprinting for ocular drug delivery, as well as both ethical and regulatory aspects, are analyzed. Through this review, we aim to raise awareness among the research community and report recent advances and future directions in order to apply this advanced therapy in the eye tissue regeneration field.

Background: Mesenchymal stem cells (MSCs) are stem cells present in adult tissues. They can be cultured, have great growth capacity, and can differentiate into several cell types. The isolation of urine-derived mesenchymal stem cells (hUSCs) was recently described. hUSCs present additional benefits in the fact that they can be easily obtained noninvasively. Regarding gene delivery, nonviral vectors based on cationic niosomes have been used and are more stable and have lower immunogenicity than viral vectors. However, their transfection efficiency is low and in need of improvement. Methods: We isolated hUSCs from urine, and the cell culture was tested and characterized. Different cationic niosomes were elaborated using reverse-phase evaporation, and they were physicochemically characterized. Then, they were screened into hUSCs for transfection efficiency, and their internalization was evaluated. Results: GPxT-CQ at a lipid/DNA ratio of 5:1 (w/w) had the best transfection efficiency. Intracellular localization studies confirmed that nioplexes entered mainly via caveolae-mediated endocytosis. Conclusions: In conclusion, we established a protocol for hUSC isolation and their transfection with cationic niosomes, which could have relevant clinical applications such as in gene therapy. This methodology could also be used for creating cellular models for studying and validating pathogenic genetic variants, and even for performing functional studies. Our study increases knowledge about the internalization of tested cationic niosomes in these previously unexplored cells

[EN]The aim was to evaluate relevant biophysic processes related to the physicochemical features and gene transfection mechanism when sphingolipids are incorporated into a cationic niosome formulation for non-viral gene delivery to central nervous system. For that, two formulations named niosphingosomes and niosomes devoid of sphingolipid extracts, as control, were developed by the oil-in water emulsion technique. Both formulations and the corresponding complexes, obtained upon the addition of the reporter EGFP plasmid, were physicochemically and biologically characterized and evaluated. Compared to niosomes, niosphingosomes, and the corresponding complexes decreased particle size and increased superficial charge. Although there were not significant differences in the cellular uptake, cell viability and transfection efficiency increased when human retinal pigment epithelial (ARPE-19) cells were exposed to niosphingoplexes. Endocytosis via caveolae decreased in the case of niosphingoplexes, which showed higher co-localization with lysosomal compartment, and endosomal escape properties. Moreover, niosphingoplexes transfected not only primary central nervous system cells, but also different cells in mouse retina, depending on the administration route, and brain cortex. These preliminary results suggest that niosphingosomes represent a promising non-viral vector formulation purposed for the treatment of both retinal and brain diseases by gene therapy approach.

26 p. : il. -- Bibliogr.: p. 24-25, Alzheimer gaixotasuna, gaixotasun neurologiko bat da, eta, dementzia da bere sintomarik nagusiena. Adinarekin agertzen den gaixotasun multifaktoriala da. Gaixotasun honen hipotesi ezagunena ß-amiloide metaketa da; baina, azken urteetan beste hainbat hipotesi aurkitzen saiatu izan dira, ß-amiloide plaka horien gain eragiten duten tratamenduen eraginaren ondorioz ikusi delako ez dela beti gaixotasunaren hobekuntza ikusten. Beraz, beste hainbat hipotesi aztertu dira, adibidez, garuneko baskularizazioaren gutxipena. Azken faktore hau azken urteetan garrantzia hartzen ari da, eta tratamendua faktore honetan oinarrituz garatzea da helburua. Gaur egun, merkatuan dauden tratamenduak sintomen gain soilik eragiten baitute eta ez gaixotasunaren garapenean. VEGF faktoreak eragin neurobabesle eta angiogenikoa dituenez Alzheimerren gaixotasunean eraginkorra izan daitekeela uste da. Horregatik, maila preklinikoan ikerketa ezberdinak egin ondoren eta horiek adierazitako emaitza positiboak aztertu ondoren, etorkizunean eraginkorra izan daitekeela uste da. Eman-bidea ere ikertzeko beste atal garrantzitsu bat izan da gaixotasun honetan, gaixotasun neurologiko bat denez ez baita horren erraza farmakoak bertara helaraztea. Kasu honetan, sudur-bidea aztertu da hesi hematoentzefalikoa gainditzeko gai izan daitekeen eman-bide berritzaile eta ez-inbaditzaile gisa, etorkizunerako aukera bat izan daitekeelakoan.

The cell-based approach in gene therapy arises as a promising strategy to provide safe, targeted, and efficient gene delivery. Owing to their unique features, as homing and tumor-tropism, mesenchymal stem cells (MSCs) have recently been introduced as an encouraging vehicle in gene therapy. Nevertheless, non-viral transfer of nucleic acids into MSCs remains limited due to various factors related to the main stakeholders of the process (e.g., nucleic acids, carriers, or cells). In this review, we have summarized the main types of nucleic acids used to transfect MSCs, the pros and cons, and applications of each. Then, we have emphasized on the most efficient lipid-based carriers for nucleic acids to MSCs, their main features, and some of their applications. While a myriad of studies have demonstrated the therapeutic potential for engineered MSCs therapy in various illnesses, optimization for clinical use is an ongoing challenge. On the way of improvement, genetically modified MSCs have been combined with various novel techniques and tools (e.g., exosomes, spheroids, 3D-Bioprinting, etc.,) aiming for more efficient and safe applications in biomedicine.

Background: Mesenchymal stem cells (MSCs) are stem cells present in adult tissues. They can be cultured, have great growth capacity, and can differentiate into several cell types. The isolation of urine-derived mesenchymal stem cells (hUSCs) was recently described. hUSCs present additional benefits in the fact that they can be easily obtained noninvasively. Regarding gene delivery, nonviral vectors based on cationic niosomes have been used and are more stable and have lower immunogenicity than viral vectors. However, their transfection efficiency is low and in need of improvement. Methods: We isolated hUSCs from urine, and the cell culture was tested and characterized. Different cationic niosomes were elaborated using reverse-phase evaporation, and they were physicochemically characterized. Then, they were screened into hUSCs for transfection efficiency, and their internalization was evaluated. Results: GPxT-CQ at a lipid/DNA ratio of 5:1 (w/w) had the best transfection efficiency. Intracellular localization studies confirmed that nioplexes entered mainly via caveolae-mediated endocytosis. Conclusions: In conclusion, we established a protocol for hUSC isolation and their transfection with cationic niosomes, which could have relevant clinical applications such as in gene therapy. This methodology could also be used for creating cellular models for studying and validating pathogenic genetic variants, and even for performing functional studies. Our study increases knowledge about the internalization of tested cationic niosomes in these previously unexplored cells

Efficient delivery of genetic material into cells is a critical process to translate gene therapy into clinical practice. In this sense, the increased knowledge acquired during past years in the molecular biology and nanotechnology fields has contributed to the development of different kinds of non-viral vector systems as a promising alternative to virus-based gene delivery counterparts. Consequently, the development of non-viral vectors has gained attention, and nowadays, gene delivery mediated by these systems is considered as the cornerstone of modern gene therapy due to relevant advantages such as low toxicity, poor immunogenicity and high packing capacity. However, despite these relevant advantages, non-viral vectors have been poorly translated into clinical success. This review addresses some critical issues that need to be considered for clinical practice application of non-viral vectors in mainstream medicine, such as efficiency, biocompatibility, long-lasting effect, route of administration, design of experimental condition or commercialization process. In addition, potential strategies for overcoming main hurdles are also addressed. Overall, this review aims to raise awareness among the scientific community and help researchers gain knowledge in the design of safe and efficient non-viral gene delivery systems for clinical applications to progress in the gene therapy field.

Lipid nanocarriers, such as niosomes, are considered attractive candidates for non-viral gene delivery due to their suitable biocompatibility and high versatility. In this work, we studied the influence of incorporating chloroquine in niosomes biophysical performance, as well as the effect of non-ionic surfactant composition and protocol of incorporation in their biophysical performance. An exhaustive comparative evaluation of three niosome formulations differing in these parameters was performed, which included the analysis of their thermal stability, rheological behavior, mean particle size, dispersity, zeta potential, morphology, membrane packing capacity, affinity to bind DNA, ability to release and protect the genetic material, buffering capacity and ability to escape from artificially synthesized lysosomes. Finally, in vitro biological studies were, also, performed in order to determine the compatibility of the formulations with biological systems, their transfection efficiency and transgene expression. Results revealed that the incorporation of chloroquine in niosome formulations improved their biophysical properties and the transfection efficiency, while the substitution of one of the non-ionic surfactants and the phase of addition resulted in less biophysical variations. Of note, the present work provides several biophysical parameters and characterization strategies that could be used as gold standard for gene therapy nanosystems evaluation.

Three-dimensional (3D) printing is a game changer technology that holds great promise for a wide variety of biomedical applications, including ophthalmology. Through this emerging technique, specific eye tissues can be custom-fabricated in a flexible and automated way, incorporating different cell types and biomaterials in precise anatomical 3D geometries. However, and despite the great progress and possibilities generated in recent years, there are still challenges to overcome that jeopardize its clinical application in regular practice. The main goal of this review is to provide an in-depth understanding of the current status and implementation of 3D bioprinting technology in the ophthalmology field in order to manufacture relevant tissues such as cornea, retina and conjunctiva. Special attention is paid to the description of the most commonly employed bioprinting methods, and the most relevant eye tissue engineering studies performed by 3D bioprinting technology at preclinical level. In addition, other relevant issues related to use of 3D bioprinting for ocular drug delivery, as well as both ethical and regulatory aspects, are analyzed. Through this review, we aim to raise awareness among the research community and report recent advances and future directions in order to apply this advanced therapy in the eye tissue regeneration field.

The cell-based approach in gene therapy arises as a promising strategy to provide safe, targeted, and efficient gene delivery. Owing to their unique features, as homing and tumor-tropism, mesenchymal stem cells (MSCs) have recently been introduced as an encouraging vehicle in gene therapy. Nevertheless, non-viral transfer of nucleic acids into MSCs remains limited due to various factors related to the main stakeholders of the process (e.g., nucleic acids, carriers, or cells). In this review, we have summarized the main types of nucleic acids used to transfect MSCs, the pros and cons, and applications of each. Then, we have emphasized on the most efficient lipid-based carriers for nucleic acids to MSCs, their main features, and some of their applications. While a myriad of studies have demonstrated the therapeutic potential for engineered MSCs therapy in various illnesses, optimization for clinical use is an ongoing challenge. On the way of improvement, genetically modified MSCs have been combined with various novel techniques and tools (e.g., exosomes, spheroids, 3D-Bioprinting, etc.,) aiming for more efficient and safe applications in biomedicine.

The main objective of three-dimensional (3D) bioprinting technology is the development of 3D constructs combining biomaterials and cells to replicate tissues and organs. Using this technique, different kinds of epitheliums can be produced in order to imitate the in vivo environment with more precision than cell cultures. Such scaffolds are usually characterized prior to evaluate cellular response upon the addition of different drugs and genetic materials. The aim of this study was to make use of 3D bioprinting technology to develop scaffolds that mimic the in vivo lung environment to evaluate the capacity of niosomes (non-viral vectors) to deliver genetic material for the treatment of cystic fibrosis (CF) genetic disorder.

The cell-based approach in gene therapy arises as a promising strategy to provide safe, targeted, and efficient gene delivery. Owing to their unique features, as homing and tumor-tropism, mesenchymal stem cells (MSCs) have recently been introduced as an encouraging vehicle in gene therapy. Nevertheless, non-viral transfer of nucleic acids into MSCs remains limited due to various factors related to the main stakeholders of the process (e.g., nucleic acids, carriers, or cells). In this review, we have summarized the main types of nucleic acids used to transfect MSCs, the pros and cons, and applications of each. Then, we have emphasized on the most efficient lipid-based carriers for nucleic acids to MSCs, their main features, and some of their applications. While a myriad of studies have demonstrated the therapeutic potential for engineered MSCs therapy in various illnesses, optimization for clinical use is an ongoing challenge. On the way of improvement, genetically modified MSCs have been combined with various novel techniques and tools (e.g., exosomes, spheroids, 3D-Bioprinting, etc.,) aiming for more efficient and safe applications in biomedicine.

[EN]The aim was to evaluate relevant biophysic processes related to the physicochemical features and gene transfection mechanism when sphingolipids are incorporated into a cationic niosome formulation for non-viral gene delivery to central nervous system. For that, two formulations named niosphingosomes and niosomes devoid of sphingolipid extracts, as control, were developed by the oil-in water emulsion technique. Both formulations and the corresponding complexes, obtained upon the addition of the reporter EGFP plasmid, were physicochemically and biologically characterized and evaluated. Compared to niosomes, niosphingosomes, and the corresponding complexes decreased particle size and increased superficial charge. Although there were not significant differences in the cellular uptake, cell viability and transfection efficiency increased when human retinal pigment epithelial (ARPE-19) cells were exposed to niosphingoplexes. Endocytosis via caveolae decreased in the case of niosphingoplexes, which showed higher co-localization with lysosomal compartment, and endosomal escape properties. Moreover, niosphingoplexes transfected not only primary central nervous system cells, but also different cells in mouse retina, depending on the administration route, and brain cortex. These preliminary results suggest that niosphingosomes represent a promising non-viral vector formulation purposed for the treatment of both retinal and brain diseases by gene therapy approach.

26 p. : il. -- Bibliogr.: p. 24-25, Alzheimer gaixotasuna, gaixotasun neurologiko bat da, eta, dementzia da bere sintomarik nagusiena. Adinarekin agertzen den gaixotasun multifaktoriala da. Gaixotasun honen hipotesi ezagunena ß-amiloide metaketa da; baina, azken urteetan beste hainbat hipotesi aurkitzen saiatu izan dira, ß-amiloide plaka horien gain eragiten duten tratamenduen eraginaren ondorioz ikusi delako ez dela beti gaixotasunaren hobekuntza ikusten. Beraz, beste hainbat hipotesi aztertu dira, adibidez, garuneko baskularizazioaren gutxipena. Azken faktore hau azken urteetan garrantzia hartzen ari da, eta tratamendua faktore honetan oinarrituz garatzea da helburua. Gaur egun, merkatuan dauden tratamenduak sintomen gain soilik eragiten baitute eta ez gaixotasunaren garapenean. VEGF faktoreak eragin neurobabesle eta angiogenikoa dituenez Alzheimerren gaixotasunean eraginkorra izan daitekeela uste da. Horregatik, maila preklinikoan ikerketa ezberdinak egin ondoren eta horiek adierazitako emaitza positiboak aztertu ondoren, etorkizunean eraginkorra izan daitekeela uste da. Eman-bidea ere ikertzeko beste atal garrantzitsu bat izan da gaixotasun honetan, gaixotasun neurologiko bat denez ez baita horren erraza farmakoak bertara helaraztea. Kasu honetan, sudur-bidea aztertu da hesi hematoentzefalikoa gainditzeko gai izan daitekeen eman-bide berritzaile eta ez-inbaditzaile gisa, etorkizunerako aukera bat izan daitekeelakoan.

Lipid nanocarriers, such as niosomes, are considered attractive candidates for non-viral gene delivery due to their suitable biocompatibility and high versatility. In this work, we studied the influence of incorporating chloroquine in niosomes biophysical performance, as well as the effect of non-ionic surfactant composition and protocol of incorporation in their biophysical performance. An exhaustive comparative evaluation of three niosome formulations differing in these parameters was performed, which included the analysis of their thermal stability, rheological behavior, mean particle size, dispersity, zeta potential, morphology, membrane packing capacity, affinity to bind DNA, ability to release and protect the genetic material, buffering capacity and ability to escape from artificially synthesized lysosomes. Finally, in vitro biological studies were, also, performed in order to determine the compatibility of the formulations with biological systems, their transfection efficiency and transgene expression. Results revealed that the incorporation of chloroquine in niosome formulations improved their biophysical properties and the transfection efficiency, while the substitution of one of the non-ionic surfactants and the phase of addition resulted in less biophysical variations. Of note, the present work provides several biophysical parameters and characterization strategies that could be used as gold standard for gene therapy nanosystems evaluation.

Background: Mesenchymal stem cells (MSCs) are stem cells present in adult tissues. They can be cultured, have great growth capacity, and can differentiate into several cell types. The isolation of urine-derived mesenchymal stem cells (hUSCs) was recently described. hUSCs present additional benefits in the fact that they can be easily obtained noninvasively. Regarding gene delivery, nonviral vectors based on cationic niosomes have been used and are more stable and have lower immunogenicity than viral vectors. However, their transfection efficiency is low and in need of improvement. Methods: We isolated hUSCs from urine, and the cell culture was tested and characterized. Different cationic niosomes were elaborated using reverse-phase evaporation, and they were physicochemically characterized. Then, they were screened into hUSCs for transfection efficiency, and their internalization was evaluated. Results: GPxT-CQ at a lipid/DNA ratio of 5:1 (w/w) had the best transfection efficiency. Intracellular localization studies confirmed that nioplexes entered mainly via caveolae-mediated endocytosis. Conclusions: In conclusion, we established a protocol for hUSC isolation and their transfection with cationic niosomes, which could have relevant clinical applications such as in gene therapy. This methodology could also be used for creating cellular models for studying and validating pathogenic genetic variants, and even for performing functional studies. Our study increases knowledge about the internalization of tested cationic niosomes in these previously unexplored cells

Three-dimensional (3D) printing is a game changer technology that holds great promise for a wide variety of biomedical applications, including ophthalmology. Through this emerging technique, specific eye tissues can be custom-fabricated in a flexible and automated way, incorporating different cell types and biomaterials in precise anatomical 3D geometries. However, and despite the great progress and possibilities generated in recent years, there are still challenges to overcome that jeopardize its clinical application in regular practice. The main goal of this review is to provide an in-depth understanding of the current status and implementation of 3D bioprinting technology in the ophthalmology field in order to manufacture relevant tissues such as cornea, retina and conjunctiva. Special attention is paid to the description of the most commonly employed bioprinting methods, and the most relevant eye tissue engineering studies performed by 3D bioprinting technology at preclinical level. In addition, other relevant issues related to use of 3D bioprinting for ocular drug delivery, as well as both ethical and regulatory aspects, are analyzed. Through this review, we aim to raise awareness among the research community and report recent advances and future directions in order to apply this advanced therapy in the eye tissue regeneration field.

Efficient delivery of genetic material into cells is a critical process to translate gene therapy into clinical practice. In this sense, the increased knowledge acquired during past years in the molecular biology and nanotechnology fields has contributed to the development of different kinds of non-viral vector systems as a promising alternative to virus-based gene delivery counterparts. Consequently, the development of non-viral vectors has gained attention, and nowadays, gene delivery mediated by these systems is considered as the cornerstone of modern gene therapy due to relevant advantages such as low toxicity, poor immunogenicity and high packing capacity. However, despite these relevant advantages, non-viral vectors have been poorly translated into clinical success. This review addresses some critical issues that need to be considered for clinical practice application of non-viral vectors in mainstream medicine, such as efficiency, biocompatibility, long-lasting effect, route of administration, design of experimental condition or commercialization process. In addition, potential strategies for overcoming main hurdles are also addressed. Overall, this review aims to raise awareness among the scientific community and help researchers gain knowledge in the design of safe and efficient non-viral gene delivery systems for clinical applications to progress in the gene therapy field.

130 p., En la tesis actual, demostramos que los nioplexos DP80 podían transfectar eficientemente célulasmesenquimales MSC D1 con los plásmidos pCMS-EGFP y pUNO1-hBMP7. Sin embargo, se requiere laoptimización del proceso de formulación de niosomas para mejorar la eficacia de la transfección y laviabilidad celular. Además, las células D1-MSC transfectadas con hBMP7 podrían considerarse no solocomo una herramienta eficaz de administración de la proteína hBMP7, sino también como célulasproliferantes y regeneradoras de hueso con fines terapéuticos, ofreciendo una esperanza razonable deutilizar esta doble estrategia basada en terapia génica mediante vectores no virales y celular mediante laadministración de células mesenquimales para la regeneración ósea. Por otro lado, nuestros resultadosmostraron que los niosomas DPP80, elaborados con una mezcla de tensioactivos no iónicos P y P80,fueron capaces de transfectar eficientemente las células NT2 sin comprometer su viabilidad. Loscomplejos DPP80-EGFP pudieron transfectar preferentemente neuronas en cultivos corticales primariosembrionarios de rata e inducir la transfección directa in vivo de células gliales en la corteza cerebral derata. Finalmente, proporcionamos una prueba de concepto de que las células NT2 podrían transfectarsecon plásmidos de expresión hBMP7 usando como vector no viral los niosomas catiónicos DPP80. Lascélulas NT2 que sobreexpresaron hBMP7 podrían dificultar la migración de las células de glioma C6.Estos resultados destacan el potencial de esta estrategia para impedir la metástasis de las células deglioma.

Non-viral vectors have emerged as a promising alternative to viral gene delivery systems due to their safer profile. Among non-viral vectors, recently, niosomes have shown favorable properties for gene delivery, including low toxicity, high stability, and easy production. The three main components of niosome formulations include a cationic lipid that is responsible for the electrostatic interactions with the negatively charged genetic material, a non-ionic surfactant that enhances the long-term stability of the niosome, and a helper component that can be added to improve its physicochemical properties and biological performance. This review is aimed at providing recent information about niosome-based non-viral vectors for gene delivery purposes. Specially, we will discuss the composition, preparation methods, physicochemical properties, and biological evaluation of niosomes and corresponding nioplexes that result from the addition of the genetic material onto their cationic surface. Next, we will focus on the in situ application of such niosomes to deliver the genetic material into immune-privileged tissues such as the brain cortex and the retina. Finally, as future perspectives, non-invasive administration routes and different targeting strategies will be discussed.

174 p., En esta tesis elaboramos y caracterizamos una nueva formulación de niosomas no virales (DP60L)basada en lípidos catiónicos DOTMA y polisorbato 60 como tensioactivo no iónico y licopeno comolípido auxiliar. Los nuevos niosomas presentaron propiedades fisicoquímicas adecuadas para aplicaciones de administración de genes en términos de tamaño, carga superficial, polidispersidad o capacidad para proteger el material genético contra la digestión enzimática. La incorporación de un compuesto natural como el licopeno en la formulación aumentó la eficiencia de transfección en las células ARPE-19, sin afectar la viabilidad celular, probablemente debido a la vía de endocitosis particular que evadió la degradación lisosómica del ADN. Las administraciones in vivo de nioplexos DP60L a la retina de rata transfectó las capas celulares externas de la retina, lo que ofrece una esperanza razonable para el tratamiento de muchas enfermedades retinianas heredadas mediante una formulación no viral segura. Porotro lado, dicha formulación fue capaz de transfectar eficientemente las células NT2 mostrando propiedades de escape endosómico que podrían explicar los altos niveles de expresión de proteínas observados. Se obtuvieron resultados prometedores en cultivos corticales primarios de embriones de rata e in vivo después de la inyección intracraneal, reforzando la aplicación futura de dichos niosomas como herramienta eficiente de entrega de genes para el tratamiento de los trastornos degenerativos y malignosdel SNC. Finalmente, la adición de otra molécula auxiliar, como la cloroquina, mejoró la transfección retiniana in vivo tras administración intravitrea y subretiniana.

130 p., En la tesis actual, demostramos que los nioplexos DP80 podían transfectar eficientemente célulasmesenquimales MSC D1 con los plásmidos pCMS-EGFP y pUNO1-hBMP7. Sin embargo, se requiere laoptimización del proceso de formulación de niosomas para mejorar la eficacia de la transfección y laviabilidad celular. Además, las células D1-MSC transfectadas con hBMP7 podrían considerarse no solocomo una herramienta eficaz de administración de la proteína hBMP7, sino también como célulasproliferantes y regeneradoras de hueso con fines terapéuticos, ofreciendo una esperanza razonable deutilizar esta doble estrategia basada en terapia génica mediante vectores no virales y celular mediante laadministración de células mesenquimales para la regeneración ósea. Por otro lado, nuestros resultadosmostraron que los niosomas DPP80, elaborados con una mezcla de tensioactivos no iónicos P y P80,fueron capaces de transfectar eficientemente las células NT2 sin comprometer su viabilidad. Loscomplejos DPP80-EGFP pudieron transfectar preferentemente neuronas en cultivos corticales primariosembrionarios de rata e inducir la transfección directa in vivo de células gliales en la corteza cerebral derata. Finalmente, proporcionamos una prueba de concepto de que las células NT2 podrían transfectarsecon plásmidos de expresión hBMP7 usando como vector no viral los niosomas catiónicos DPP80. Lascélulas NT2 que sobreexpresaron hBMP7 podrían dificultar la migración de las células de glioma C6.Estos resultados destacan el potencial de esta estrategia para impedir la metástasis de las células deglioma.

Non-viral vectors have emerged as a promising alternative to viral gene delivery systems due to their safer profile. Among non-viral vectors, recently, niosomes have shown favorable properties for gene delivery, including low toxicity, high stability, and easy production. The three main components of niosome formulations include a cationic lipid that is responsible for the electrostatic interactions with the negatively charged genetic material, a non-ionic surfactant that enhances the long-term stability of the niosome, and a helper component that can be added to improve its physicochemical properties and biological performance. This review is aimed at providing recent information about niosome-based non-viral vectors for gene delivery purposes. Specially, we will discuss the composition, preparation methods, physicochemical properties, and biological evaluation of niosomes and corresponding nioplexes that result from the addition of the genetic material onto their cationic surface. Next, we will focus on the in situ application of such niosomes to deliver the genetic material into immune-privileged tissues such as the brain cortex and the retina. Finally, as future perspectives, non-invasive administration routes and different targeting strategies will be discussed.

174 p., En esta tesis elaboramos y caracterizamos una nueva formulación de niosomas no virales (DP60L)basada en lípidos catiónicos DOTMA y polisorbato 60 como tensioactivo no iónico y licopeno comolípido auxiliar. Los nuevos niosomas presentaron propiedades fisicoquímicas adecuadas para aplicaciones de administración de genes en términos de tamaño, carga superficial, polidispersidad o capacidad para proteger el material genético contra la digestión enzimática. La incorporación de un compuesto natural como el licopeno en la formulación aumentó la eficiencia de transfección en las células ARPE-19, sin afectar la viabilidad celular, probablemente debido a la vía de endocitosis particular que evadió la degradación lisosómica del ADN. Las administraciones in vivo de nioplexos DP60L a la retina de rata transfectó las capas celulares externas de la retina, lo que ofrece una esperanza razonable para el tratamiento de muchas enfermedades retinianas heredadas mediante una formulación no viral segura. Porotro lado, dicha formulación fue capaz de transfectar eficientemente las células NT2 mostrando propiedades de escape endosómico que podrían explicar los altos niveles de expresión de proteínas observados. Se obtuvieron resultados prometedores en cultivos corticales primarios de embriones de rata e in vivo después de la inyección intracraneal, reforzando la aplicación futura de dichos niosomas como herramienta eficiente de entrega de genes para el tratamiento de los trastornos degenerativos y malignosdel SNC. Finalmente, la adición de otra molécula auxiliar, como la cloroquina, mejoró la transfección retiniana in vivo tras administración intravitrea y subretiniana.

130 p., En la tesis actual, demostramos que los nioplexos DP80 podían transfectar eficientemente célulasmesenquimales MSC D1 con los plásmidos pCMS-EGFP y pUNO1-hBMP7. Sin embargo, se requiere laoptimización del proceso de formulación de niosomas para mejorar la eficacia de la transfección y laviabilidad celular. Además, las células D1-MSC transfectadas con hBMP7 podrían considerarse no solocomo una herramienta eficaz de administración de la proteína hBMP7, sino también como célulasproliferantes y regeneradoras de hueso con fines terapéuticos, ofreciendo una esperanza razonable deutilizar esta doble estrategia basada en terapia génica mediante vectores no virales y celular mediante laadministración de células mesenquimales para la regeneración ósea. Por otro lado, nuestros resultadosmostraron que los niosomas DPP80, elaborados con una mezcla de tensioactivos no iónicos P y P80,fueron capaces de transfectar eficientemente las células NT2 sin comprometer su viabilidad. Loscomplejos DPP80-EGFP pudieron transfectar preferentemente neuronas en cultivos corticales primariosembrionarios de rata e inducir la transfección directa in vivo de células gliales en la corteza cerebral derata. Finalmente, proporcionamos una prueba de concepto de que las células NT2 podrían transfectarsecon plásmidos de expresión hBMP7 usando como vector no viral los niosomas catiónicos DPP80. Lascélulas NT2 que sobreexpresaron hBMP7 podrían dificultar la migración de las células de glioma C6.Estos resultados destacan el potencial de esta estrategia para impedir la metástasis de las células deglioma.

Non-viral vectors have emerged as a promising alternative to viral gene delivery systems due to their safer profile. Among non-viral vectors, recently, niosomes have shown favorable properties for gene delivery, including low toxicity, high stability, and easy production. The three main components of niosome formulations include a cationic lipid that is responsible for the electrostatic interactions with the negatively charged genetic material, a non-ionic surfactant that enhances the long-term stability of the niosome, and a helper component that can be added to improve its physicochemical properties and biological performance. This review is aimed at providing recent information about niosome-based non-viral vectors for gene delivery purposes. Specially, we will discuss the composition, preparation methods, physicochemical properties, and biological evaluation of niosomes and corresponding nioplexes that result from the addition of the genetic material onto their cationic surface. Next, we will focus on the in situ application of such niosomes to deliver the genetic material into immune-privileged tissues such as the brain cortex and the retina. Finally, as future perspectives, non-invasive administration routes and different targeting strategies will be discussed.

Cationic niosomes have become important non-viral vehicles for transporting a good number of small drug molecules and macromolecules. Growing interest shown by these colloidal nanoparticles in therapy is determined by their structural similarities to liposomes. Cationic niosomes are usually obtained from the self-assembly of non-ionic surfactant molecules. This process can be governed not only by the nature of such surfactants but also by others factors like the presence of additives, formulation preparation and properties of the encapsulated hydrophobic or hydrophilic molecules. This review is aimed at providing recent information for using cationic niosomes for gene delivery purposes with particular emphasis on improving the transportation of antisense oligonucleotides (ASOs), small interference RNAs (siRNAs), aptamers and plasmids (pDNA).

Cationic niosomes have become important non-viral vehicles for transporting a good number of small drug molecules and macromolecules. Growing interest shown by these colloidal nanoparticles in therapy is determined by their structural similarities to liposomes. Cationic niosomes are usually obtained from the self-assembly of non-ionic surfactant molecules. This process can be governed not only by the nature of such surfactants but also by others factors like the presence of additives, formulation preparation and properties of the encapsulated hydrophobic or hydrophilic molecules. This review is aimed at providing recent information for using cationic niosomes for gene delivery purposes with particular emphasis on improving the transportation of antisense oligonucleotides (ASOs), small interference RNAs (siRNAs), aptamers and plasmids (pDNA).

Cationic niosomes have become important non-viral vehicles for transporting a good number of small drug molecules and macromolecules. Growing interest shown by these colloidal nanoparticles in therapy is determined by their structural similarities to liposomes. Cationic niosomes are usually obtained from the self-assembly of non-ionic surfactant molecules. This process can be governed not only by the nature of such surfactants but also by others factors like the presence of additives, formulation preparation and properties of the encapsulated hydrophobic or hydrophilic molecules. This review is aimed at providing recent information for using cationic niosomes for gene delivery purposes with particular emphasis on improving the transportation of antisense oligonucleotides (ASOs), small interference RNAs (siRNAs), aptamers and plasmids (pDNA).

Duración (en horas): Más de 50 horas Destinatario: Estudiante, En esta asignatura, el alumnado deberá aplicar diferentes conceptos necesarios para gestionar eficientemente la oficina de farmacia. Para ello, trabajará diferentes conceptos relaccionados con el marketing, la contabilidad, el control de stock, la gestión de compras y de ventas, etc. Todo ello, con la finalidad de conseguir la máxima rentabilidad posible en un establecimiento de interés sanitario público y gestión privada como es la oficina de farmacia, respetando siempre las normas y leyes que regulan su funcionamiento. Para el desarrollo de la mayor parte del programa se utilizarán metodologías activas, principalmente Aprendizaje Basado en Problemas (ABP), aunque también se llevarán a cabo otras actividades colaborativas como por ejemplo el aprendizaje a través del Puzzle de Aronson o GPS (Gided Problem Solving) cuyas carcaterísticas diferenciales se irán explicando en el momento de aplicarlas en el aula. Con el ABP lo que se pretende es que el alumnado sea capaz, a través de una situación inicial cercana a lo que le ocurrirá en su futuro profesional, de pensar qué información necesita para poder hacer frente con éxito a ese probelma o situación. Es decir, el alumnado debe identificar las necesidades u objetivos de aprendizaje. Una vez identificados, será necesario un trabajo de búsqueda e interiorización de información, así como de su aplicación a ciertas situaciones de una manera autónoma, aunque siempre guiada por el profesorado, que facilitará el aprendizaje continuado del alumnado. Al contrario que en el desarrollo de la actividad de una manera más tradicional, en esta metodología, el profesorado no explica una teoría que después se ha de aplicar, sino que es el alumnado el que ha de llegar a buscar y comprender la teoría que antes se explicaba en las clases magistrales para poder resolver una situación hasta ese momento desconocida.

Duración (en horas): Más de 50 horas Destinatario: Estudiante, En esta asignatura, el alumnado deberá aplicar diferentes conceptos necesarios para gestionar eficientemente la oficina de farmacia. Para ello, trabajará diferentes conceptos relaccionados con el marketing, la contabilidad, el control de stock, la gestión de compras y de ventas, etc. Todo ello, con la finalidad de conseguir la máxima rentabilidad posible en un establecimiento de interés sanitario público y gestión privada como es la oficina de farmacia, respetando siempre las normas y leyes que regulan su funcionamiento. Para el desarrollo de la mayor parte del programa se utilizarán metodologías activas, principalmente Aprendizaje Basado en Problemas (ABP), aunque también se llevarán a cabo otras actividades colaborativas como por ejemplo el aprendizaje a través del Puzzle de Aronson o GPS (Gided Problem Solving) cuyas carcaterísticas diferenciales se irán explicando en el momento de aplicarlas en el aula. Con el ABP lo que se pretende es que el alumnado sea capaz, a través de una situación inicial cercana a lo que le ocurrirá en su futuro profesional, de pensar qué información necesita para poder hacer frente con éxito a ese probelma o situación. Es decir, el alumnado debe identificar las necesidades u objetivos de aprendizaje. Una vez identificados, será necesario un trabajo de búsqueda e interiorización de información, así como de su aplicación a ciertas situaciones de una manera autónoma, aunque siempre guiada por el profesorado, que facilitará el aprendizaje continuado del alumnado. Al contrario que en el desarrollo de la actividad de una manera más tradicional, en esta metodología, el profesorado no explica una teoría que después se ha de aplicar, sino que es el alumnado el que ha de llegar a buscar y comprender la teoría que antes se explicaba en las clases magistrales para poder resolver una situación hasta ese momento desconocida.

The following article presents the experience of a teaching innovation introduced in the subject of Management, Planning, Legislation and Deontology of the Degree in Pharmacy of the University of the Basque Country UPV/EHU, to learn the thematic block of Marketing and Merchandising. This educational action has focused on achieving both the specific competences related to this subject, as well as transversal competences of team work, information search and reflection capacity, among others. For this purpose, the Problem-Based Learning (PBL) methodology has been used, making the student to face situations that will happen to him in his professional future as a pharmacist. Interactive questionnaires designed by rapid response devices (clickers), video visualizations, expository activities and reports were used. The students sometimes worked autonomously, but mainly the work in teams was promoted where the work done by each student was essential for the resolution of the situation. The valuation, both formative and accumulative, was carried out through a group deliverable, an individual self-assessment test, two brief group presentations, two individual assessment tests and finally, a more extensive group presentation. In general the evaluation showed highly satisfactory results. In addition, great involvement was achieved by the students making the development of the classes much more active and dynamic., En el siguiente artículo se presenta la experiencia de innovación docente introducida en la asignatura de Gestión, Planificación, Legislación y Deontología del Grado en Farmacia de la Universidad del País Vasco UPV/EHU, para el aprendizaje del bloque temático de Marketing y Merchandising. Esta acción educativa se ha enfocado en alcanzar tanto las competencias específicas relacionadas con este tema, así como las competencias transversales de trabajo en equipo, búsqueda de información y capacidad de reflexión, entre otras. Para ellos se ha utilizado la metodología de Aprendizaje Basado en Problemas (ABP) haciendo que el estudiante se encuentre ante situaciones que le acontecerán en su futuro profesional como farmacéutico. Se utilizaron cuestionarios interactivos diseñados con dispositivos de respuesta rápida (clickers), visualizaciones de videos, exposiciones e informes. Los alumnos trabajaron en ocasiones de manera autónoma, pero principalmente se impulsó el trabajo en equipos donde el trabajo realizado por cada componente del mismo fuera esencial para la resolución de la situación planteada. La evaluación tanto formativa como sumativa, se realizó por medio de un entregable grupal, un test de autoevaluación individual, dos presentaciones breves grupales, dos test de evaluación individual y una presentación grupal, algo más extensa. En general, los resultados obtenidos fueron altamente satisfactorios. Además, se logró una gran implicación por parte del alumnado haciendo el desarrollo de las clases mucho más activo y dinámico.