Novel PEI-aldehyde conjugates for gene delivery: Promoting chondrogenic differentiation in human mesenchymal stem cells

Mesenchymal stem cell (MSC) gene therapy holds significant potential for regenerative medicine, especially for treating conditions such as cartilage damage. Still, finding appropriate vectors to achieve a safe and efficient gene delivery remains a challenge. This study explores the development of novel polyethyleneimine (PEI)-based polymers functionalized with both cationic guanidinium and hydrophobic aldehyde groups for efficient transfection to human MSCs (hMSCs). PEI was chemically modified with guanidinium-(3-guanidin-N-(3-oxopropyl)propanamide [T1]) and 1-(4-formylphenyl)guanidine [T2]) and hydrophobic (octanal [T3A] and dodecanal [T3B]) aldehydes. Polyplexes were formed by the complexation of PEI-aldehyde conjugates with plasmids encoding for β-galactosidase (p<i>lacZ</i>), green fluorescent protein (pGFP), and the chondrogenic transcription factor SOX9 (p<i>sox9</i>), and demonstrated efficient DNA complexation and protection. Among the formulations, PEI functionalized with the cationic (T2) and hydrophobic (T3A) aldehydes (PEIT2T3A) exhibited a superior transfection efficiency and biocompatibility, significantly enhancing the expression of target genes in hMSCs. Importantly, PEIT2T3A/p<i>sox9</i> polyplexes successfully promoted the chondrogenic differentiation of hMSCs, as evidenced by the increased expression of chondrogenic markers (SOX9, type-II collagen [COLII], and aggrecan [ACAN]) and proteoglycan deposition in aggregate cultures, while mitigating the low cell viability found with unmodified PEI. These findings suggest that PEIT2T3A is a promising non-viral vector for targeted gene delivery and hMSC-based regenerative medicine applications.