Physical and mechanical cues affecting biomaterial-mediated plasmid DNA delivery: insights into non-viral delivery systems

Abstract

Background Whilst traditional strategies to increase transfection efficiency of non-viral systems aimed at modifying the vector or the polyplexes/lipoplexes, biomaterial-mediated gene delivery has recently sparked increased interest. This review aims at discussing biomaterial properties and unravelling underlying mechanisms of action, for biomaterial-mediated gene delivery. DNA internalisation and cytoplasmic transport are initially discussed. DNA immobilisation, encapsulation and surface-mediated gene delivery (SMD), the role of extracellular matrix (ECM) and topographical cues, biomaterial stiffness and mechanical stimulation are finally outlined. Main text Endocytic pathways and mechanisms to escape the lysosomal network are highly variable. They depend on cell and DNA complex types but can be diverted using appropriate biomaterials. 3D scaffolds are generally fabricated via DNA immobilisation or encapsulation. Degradation rate and interaction with the vector affect temporal patterns of DNA release and transgene expression. In SMD, DNA is instead coated on 2D surfaces. SMD allows the incorporation of topographical cues, which, by inducing cytoskeletal re-arrangements, modulate DNA endocytosis. Incorporation of ECM mimetics allows cell type-specific transfection, whereas in spite of discordances in terms of optimal loading regimens, it is recognised that mechanical loading facilitates gene transfection. Finally, stiffer 2D substrates enhance DNA internalisation, whereas in 3D scaffolds, the role of stiffness is still dubious. Conclusion Although it is recognised that biomaterials allow the creation of tailored non-viral gene delivery systems, there still are many outstanding questions. A better characterisation of endocytic pathways would allow the diversion of cell adhesion processes and cytoskeletal dynamics, in order to increase cellular transfection. Further research on optimal biomaterial mechanical properties, cell ligand density and loading regimens is limited by the fact that such parameters influence a plethora of other different processes (e.g. cellular adhesion, spreading, migration, infiltration, and proliferation, DNA diffusion and release) which may in turn modulate gene delivery. Only a better understanding of these processes may allow the creation of novel robust engineered systems, potentially opening up a whole new area of biomaterial-guided gene delivery for non-viral systems.

Bioengineered Hierarchical Bonelike Compartmentalized Microconstructs Using Nanogrooved Microdiscs

“…Additionally, adhesion-associated proteins like ITGB1 were less abundant in MVs secreted from cells grown on topodiscs. Accordingly, other integrins were reported to be downregulated in MSCs cultured on nanopatterned ridges, and nanoscale surface topography was observed by atomic force microscopy to reduce focal adhesions and enhance integrin endocytosis on various cells . Although apparently counter-intuitive, for cells to be able to undergo the profound cytoskeleton and membranar alterations associated with differentiation, their initial morphology must change, including initial F-actin stress fiber dismantling and some degree of cell detachment from the substrate. , Taken together, the results of this pilot assay strengthen the impact of nanogrooved topography in dynamic coculturing and highlight the importance to perform a deeper and comprehensive analysis on the effects of topography on each cell type (ASC and HUVEC) intracellular processes, mechanisms, and signaling, particularly the ones involved in key processes emphasized here like proliferation, adhesion, and signaling by Wnt, Notch, and HH.…”

Development of a fibrin-mediated gene delivery system for the treatment of cystinosis via design of experiment

“…In all subsequent experiments, the K2 reagent, at a 1:3 DNA: reagent ratio was used. DNA-containing scaffolds are typically fabricated through either encapsulation or immobilisation 8 . Thus—as indicated in the sequence diagram in Fig.…”

“…As a matter of fact, fabrication parameters differently regulate a plethora of processes, such as DNA diffusion and release, biomaterial stiffness and degradation, cellular ability to infiltrate, migrate and proliferate. All these processes in turn affect DNA uptake and transgene expression 8 . For instance, in vivo biomaterials are required to degrade as rapidly as possible, for safety/cytotoxicity concerns.…”

“…Among new therapeutic approaches, biomaterial-mediated gene therapy potentially represents a cure for cystinosis. This approach would reduce side effects and safety concerns associated with systemic gene therapy, and it would treat difficulty accessible organs, such as the cornea 8 . Fibrin hydrogels appear good candidates, as they possess therapeutic potential alone, and are already used in the form of glue 9 , sealant 10 , or scaffolds to expand cells 11 for clinical applications.…”

“…Yet, a higher rate of degradation was observed in gels with no aprotinin, compared to gels containing 50 µg/mL aprotinin 19 . However, comprehensive studies of the interaction between different parameters are missing 8 . This study aimed at developing fibrin hydrogels releasing a non-viral plasmid encoding for CTNS, which can potentially be used for the treatment of cystinosis.…”