Delivery of Bioactive Gene Particles via Gelatin-Collagen-PEG-Based Electrospun Matrices

Abstract

The fabrication of fiber mats via electrospinning has been adopted in the last decades to produce high quality scaffolds for tissue engineering. However, an effective combination of electrospinning methods with gene delivery therapies remains a challenge. In this study, we describe how the delivery of gene complexes via electrospun mats that contain different volumes of gelatin (Gel), collagen (Col), and polyethylene glycol (PEG) can affect gene expression by transfected cells. Non-viral complexes were formulated by using lipid modified polyethylenimine (PEI) polymer and plasmid DNAs (pDNA) like the reporter Green Fluorescent Protein (GFP) and the therapeutically relevant Bone Morphogenetic Protein-2 (BMP-2) and electrospuned after being mixed with different volumes of Gel-Col-PEG mats and delivered to human myoblast (C2C12) and mouse osteoblast cells (MC3T3). The entrapment of GFP complexes via different homogeneous electrospun fiber mats revealed that a high fraction of collagen in the mats affected the quality of the fibers and led to reduced transfection efficiency on target cells. On the other hand, the fabrication of double-layered mats that contained collagen without complexes as a first layer and gelatin-collagen-PEG with complexes as a second layer successfully induced GFP expression and ALP activity in C2C12 cells. We conclude that this study has established the advantage of formulating multilayered bioactive collagen-based mats for gene delivery applications.

A Polyplex in a Shell: The Effect of Poly(aspartic acid)-Mediated Calcium Carbonate Mineralization on Polyplexes Properties and Transfection Efficiency

“…Therefore, the heparin-triggered dissociation of polyplexes could be an important tool to evaluate which polyplex formulations have potential to become efficient transfection agents when phenomena leading to increased robustness like the ones discussed here are in question. Notably, we were able to readily obtain transfection with the unmineralized polyplexes before (unlike this study), but these studies employed polyplexes formed in tissue culture medium (with Ca 2+ present) for a short duration of time (30 min complexation vs 120 min here). It seems like robustness is the effect allowing the transfection efficiency to happen, and the calcium excess modulates transfection efficiency, since samples with intermediate robustness (calcium-incubated samples except P1 incubated with 20 mM CaCl 2 ) did not show intermediate transfection efficiency.…”

“…The protective effect of the mineral shell could also prove to be useful in tissue engineering applications using gene-activated matrices carrying polyplexes. In electrospinning, for example, polyplexes inside a spun fiber are subjected to several conditions that could reduce transfection efficiency, such as solvents, shear forces, temperature variation, and solvent drying inside a polymer matrix. , In this scenario, mineralized polyplexes are more likely to maintain efficiency until the gene activated matrix is employed.…”

Electrospun-Fibrous-Architecture-Mediated Non-Viral Gene Therapy Drug Delivery in Regenerative Medicine

“…Considering that this design principle allows high versatility in the construction of fibrous architectures with physico-chemical, morphological and biological features that can be easily modulated to better suit the mechanical, biological and physical demand of the targeted host tissue, this thriving approach is of particular importance to regenerative medicine. Therefore, numerous blends consisting of synthetic and natural polymers in different formulations (e.g., PCL/alginate [96], PLLA/Collagen [118], PCL/silk fibroin [120], Gelatin/Collagen/PEG [144], PLLA/silk fibroin [145]) were constructed and investigated as electrospun fibrous strategies for the efficient delivery of genetic material. For instance, Zhao et al [118] managed to substantially improve transfection efficiency and rhBMP-2 transgene mRNA expression as well as to induce ectopic bone formation both in vitro and in vivo by adsorbing rhBMP-2/p-DNA complexes onto PLLA/Collagen I electrospun scaffolds.…”

“…The proposed therapy determined the enhancement of cell migration and the tubular-forming capacity of HMEC-1 cells cultured in vitro, along with a significant increase in angiogenesis compared to controls, which were tested in vivo in a full-thickness skin wounding model in C57BL/6J mice. Recently, instead of classical mono-layer mats for gene delivery applications, Tsekoura and co-workers [144] took a different approach by constructing a two-layered mat consisting of collagen fibers in the first layer and a mixture of gelatin, collagen and PEGembedding pDNA complexes in the second layer, which showed a great ability to increase the transgene expression and induce osteogenic activity in vitro, sustaining their potential for further in vivo/clinical investigations.…”

siRNA Functionalized Lipid Nanoparticles (LNPs) in Management of Diseases

“…PEG, a non-ionic biocompatible synthetic polymer, is soluble both in aqueous and non-aqueous solvents [68]. Incorporation of PEG provides the LNPs with an external polymeric layer onto their outer shell, and this helps to hinder the adsorption of serum proteins and the components of a phagocytic system, thereby extending their in vivo circulation time.…”