Commitment of human mesenchymal stromal cells towards ACL fibroblast differentiation upon rAAV-mediated FGF-2 and TGF-β overexpression using pNaSS-grafted PCL films

Abstract

Despite various clinical options, human anterior cruciate ligament (ACL) lesions do not fully heal. Biomaterial‐guided gene therapy using recombinant adeno‐associated virus (rAAV) vectors may improve the intrinsic mechanisms of ACL repair. Here, we examined whether poly(sodium styrene sulfonate)‐grafted poly(ε‐caprolactone) (pNaSS‐grafted PCL) films can deliver rAAV vectors coding for the reparative basic fibroblast growth factor (FGF‐2) and transforming growth factor beta (TGF‐β) in human mesenchymal stromal cells (hMSCs) as a source of implantable cells in ACL lesions. Efficient and sustained rAAV‐mediated reporter (red fluorescent protein) and therapeutic (FGF‐2 and TGF‐β) gene overexpression was achieved in the cells for at least 21 days in particular with pNaSS‐grafted PCL films relative to all other conditions (up to 5.2‐fold difference). Expression of FGF‐2 and TGF‐β mediated by rAAV using PCL films increased the levels of cell proliferation, the DNA contents, and the deposition of proteoglycans and of type‐I and ‐III collagen (up to 2.9‐fold difference) over time in the cells with higher levels of transcription factor expression (Mohawk, Scleraxis) (up to 1.9‐fold difference), without activation of inflammatory tumor necrosis alpha especially when using pNaSS‐grafted PCL films compared with the controls. Overall, the effects mediated by TGF‐β were higher than those promoted by FGF‐2, possibly due to higher levels of gene expression achieved upon rAAV gene transfer. This study shows the potential of using functionalized PCL films to apply rAAV vectors for ACL repair.