Prolonged Three-Dimensional Co-Delivery of Yamanaka Factors for Cell Reprogramming

Abstract

Reprogramming somatic cells into a pluripotent state has been widely investigated in two-dimensional (2D) systems but not described in the more biologically faithful three-dimensional (3D) scaffolds. Here, we devise a 3D porous tissue engineering scaffold that could achieve successful and efficient induction of pluripotency. To construct this 3D scaffold, nonviral hybrid nanoparticles were fabricated beforehand by employing calcium phosphate and cationized Pleurotus eryngii polysaccharide to codeliver plasmids OCT4, SOX2, KLF4 ,and C-MYC (pOSKM). These hybrid nanoparticles were then loaded into a 3D porous collagen scaffold to obtain the so-called pOSKM-activated 3D scaffold. This 3D scaffold could reprogram human umbilical cord mesenchymal stem cells (HUMSCs) into a pluripotent state, generating 3D cell spheres which showed positive expression of pluripotency markers in the 3D scaffolds and tightly packed colonies when transferred to 2D feeder layers. Besides sharing similar morphology, epigenetic modification, and expression of pluripotency genes with the embryonic stem cells, the 3D system-generated colonies could also be expanded on feeder layers for more than 20 passages, indicating the successful establishment of stable induced pluripotent stem cell (iPSC) lines. Our findings represent a first employment of porous 3D scaffolds to achieve successful reprogramming via a one-time transfection, offering a safe, simple, and effective alternative strategy for iPSC generation.

Toward Customized Extracellular Niche Engineering: Progress in Cell‐Entrapment Technologies

“…In a recent study, PEG hydrogel decorated with ECM peptides including adhesion peptide RGDSP was demonstrated to be a better mimic of 3D microenvironment to maintain higher pluripotency and reprogramming efficiency of iPSCs . Although it was interesting to note that the PEG hydrogel without the enrichment peptides led to comparable reprogramming of cells as that of 3D Matrigel or on a 3D collagen scaffold . User‐defined control in synthesis of such designer sECMs rely on “click” chemistries that form orthogonal mesh‐like polymeric architecture, thus the functional cues incorporated are also orthogonally arranged, unlike uniquely sequestered cues within the ECM macromolecule in tissues.…”

Engineering Biomaterials with Micro/Nanotechnologies for Cell Reprogramming

“…Interestingly, the hiPSC reprogramming can be accelerated upon incorporating calcium-phosphate-based hybrid nanoparticles into 3D collagen scaffolds, benefiting from the controlled release of OSKM genes and the biophysical regulation of the 3D niche. 193 Additionally, gold nanoparticles have emerged as another promising platform for gene and drug delivery due to their high surface area, inertness, and easy fabrication. 194 Cationic gold nanoparticles (AuNPs) are efficient carriers because they can interact with nucleic acids as well as the negatively charged plasma membrane.…”

“…The delivered nucleic acids formed ionic complexes with cationic calcium ions through their electrostatic interactions, which were easily transported into cells via ion-channel-mediated endocytosis. Interestingly, the hiPSC reprogramming can be accelerated upon incorporating calcium-phosphate-based hybrid nanoparticles into 3D collagen scaffolds, benefiting from the controlled release of OSKM genes and the biophysical regulation of the 3D niche . Additionally, gold nanoparticles have emerged as another promising platform for gene and drug delivery due to their high surface area, inertness, and easy fabrication .…”

Gold Nanoparticle Promoted Formation and Biological Properties of Injectable Hydrogels

“…Injectable hydrogels show great promise in controlled delivery of therapeutics, − cell delivery, − tissue culture, − and wound healing process. − Crucial requirements for a successful injectable hydrogel system are (i) low viscosity of precursor solution for facile injection and low heat generation during hydrogel formation, (ii) rapid gelation under the physiological condition to minimize the release of encapsulated guest molecules and spreading of solution, (iii) degradability to facilitate transport of nutrients, and (iv) cytocompatibility of hydrogels and the species formed by the biodegradation of the hydrogels. , Required properties of an injectable hydrogel should be manipulated on the basis of type of application. For example, mechanical property of an injectable hydrogel should be higher than that of a tissue to be cultured. ,, For drug delivery application, the prepolymer should be able to load hydrophilic and hydrophobic drugs, and the hydrogel should be able to release the molecules in a sustained manner.…”