Spontaneous gene transfection of human bone cells using 3D mineralized alginate–chitosan macrocapsules

Abstract

The effectiveness of nonviral gene therapy remains uncertain because of low transfection efficiencies and high toxicities compared with viral-based strategies. We describe a simple system for transient transfection of continuous human cell lines, with low toxicity, using mineral-coated chitosan and alginate capsules. As proof-of-concept, we demonstrate transfection of Saos-2 and MG63 human osteosarcoma continuous cell lines with gfp, LacZ reporter genes, and a Sox-9 carrying plasmid, to illustrate expression of a functional gene with therapeutic relevance. We show that continuous cell lines transfect with significant efficiency of up to 65% possibly through the interplay between chitosan and DNA complexation and calcium/phosphate-induced translocation into cells entrapped within the 3D polysaccharide based environment, as evidenced by an absence of transfection in unmineralized and chitosan-free capsules. We demonstrated that our transfection system was equally effective at transfection of primary human bone marrow stromal cells. To illustrate, the Sox-9, DNA plasmid was spontaneously expressed in primary human bone marrow stromal cells at 7 days with up to 90% efficiency in two repeats. Mineralized polysaccharide macrocapsules are gene delivery vehicles with a number of biological and practical advantages. They are highly efficient at self-transfecting primary bone cells, with programmable spatial and temporal delivery prospects, premineralized bone-like environments, and have no cytotoxic effects, as compared with many other nonviral systems.

In vitro three-dimensional cell cultures for bone sarcomas

“…In addition to its role in anti-tumour drug resistance, TME plays a role in cell accessibility for genetic manipulation. 3D mineralized alginate-chitosan cell encapsulation resulted in an efficient tool for gene transfection in human bone cells [79] . Polysaccharide beads facilitated gene uptake by SaOS2 cells when specific calcium phosphate and chitosan rate were used indicating that microcapsule environment composition is crucial for gene transfection in 3D bone model [79] .…”

From Spheroids to Bioprinting: A Literature Review on Biomanufacturing strategies of 3D In vitro Osteosarcoma Models

“…[98] Chitosan has a similar structure to GAGs, like alginate, and promotes cell adhesion and proliferation, having been used in bone tissue engineering. [99] More recently, a few chitosan scaffolds for culturing OS cells have been reported, typically in combination with other materials, such as alginate [100] and HAp [101] , in order to enhance its poor mechanical properties. [102] Decellularized Extracellular Matrix : The ECM produced by cells present in the bone TME has been used to make 3D scaffolds for bone regeneration and in vitro tumor models, due to its structural and compositional biomimicry of the bone ECM.…”