Engineering Endostatin-Producing Cartilaginous Constructs for Cartilage Repair Using Nonviral Transfection of Chondrocyte-Seeded and Mesenchymal-Stem-Cell-Seeded Collagen Scaffolds

Abstract

Although there is widespread recognition of the importance of angiogenesis in tissue repair, there is little work on the inhibition of angiogenesis in the context of tissue engineering of naturally avascular tissues, like articular cartilage. The objective was to engineer a collagen-scaffold-based cartilaginous construct overexpressing a potent antiangiogenic factor, endostatin, using nonviral transfection. Endostatin-plasmid-supplemented collagen scaffolds were seeded with mesenchymal stem cells and chondrocytes and cultured for 20–22 days. The effects of the following variables on endostatin expression and chondrogenesis were examined: collagen scaffold material, method of nonviral vector incorporation, plasmid load, culture medium, and oxygen tension. An increase and peak of endostatin protein was observed during the first week of culture, followed by a decrease to low levels, suggesting that overexpression of endostatin could be sustained for several days using the nonviral vector. The amount of endostatin produced was tunable with the external factors. Chondrogenesis was observed in the engineered constructs cultured in chondrogenic medium at the 3-week time point, demonstrating that endostatin did not inhibit the chondrogenic potential of mesenchymal stem cells or the general viability of the cells. The ability to engineer endostatin-expressing cartilaginous constructs will be of value for future work exercising regulatory control of angiogenesis in cartilage repair.

Dermatan sulphate in methoxy polyethylene glycol-polylactide-co-glycolic acid scaffolds upregulates fibronectin gene expression but has no effect on in vivo osteochondral repair

“…This study illustrates that optimising in vitro properties of repair scaffolds can be insufficient to gain an improved repair response in vivo. Small changes in vitro might easily be overridden in vivo by much more potent factors present in the challenging environment in which cartilage repair is expected to take place such as hypoxia and mechanical loading of the repair area [31,32], which might be dealt with by improving the mechanical stability of the scaffolds and controlling the intended hypoxic environment [33,34]. The repair model being an osteochondral drill hole defect has limitations in comparison to an isolated cartilage defect where the subchondral bone plate is left intact.…”

Strategies for controlled delivery of biologics for cartilage repair

“…As a result, there has been interest in introducing anti-angiogenic factors to inhibit blood vessel growth and restore cartilage tissue to its natural state of avascularity [51]. This class of factors includes endostatin [52–54], suramin [55, 56], Flt-1 [57, 58], and bevacizumab [59]. While these factors demonstrate the ability to block vascularization and inhibit the activity of vascular endothelial growth factors (VEGFs), a clear benefit of this approach in a cartilage defect model remains to be seen.…”

Genetic engineering of mesenchymal stem cells by non-viral gene delivery

“…And the DNA incorporated in the scaffolds is capable of long-term in vivo transfection. Therefore, reverse transfection offers high potential in tissue engineering and regenerative medicine, especially for to bone regeneration [30,70,121,160] and cartilage repair [62,77].…”

“…Up to this date, several natural and synthesized substrate materials have been utilized for reverse transfection of MSCs including gelatin [84], collagen [13,70,77], fibronectin [171], poly (ethylene glycol) [96], PLGA [73] and poly-L-lactic acid) [80].…”