Tissue-Engineered Cartilage Constructs Using Composite Hyaluronic Acid/Collagen I Hydrogels and Designed Poly(Propylene Fumarate) Scaffolds

Abstract

Our approach to cartilage tissue-engineering scaffolds combines image-based design and solid free-form (SFF) fabrication to create load-bearing constructs with user-defined parameters. In this study, 3-dimensional scaffolds with cubic and ellipsoidal pore architecture were fabricated using poly(propylene fumarate) (PPF). To increase seeding efficiency and cellular retention, hydrogels were used to deliver cells into the scaffolds. The first objective of this study was to evaluate the concentrations of composite hyaluronic acid (HyA) and collagen I hydrogels best able to stimulate proteoglycan synthesis in porcine chondrocytes in vitro and in vivo. The second objective was to evaluate the differences in extracellular matrix production due to pore geometry and scaffold design. For the in vitro assessment, chondrocytes were encapsulated in collagen I hydrogels with varying concentrations of HyA. Hydrogels were cultured for 1 and 2 weeks, and then the sulfated glycosaminoglycan (sGAG) content was quantified using a dimethyl-methylene blue assay. The concentration of HyA best able to increase ECM synthesis was 5% HyA/collagen I, or 0.23 mg/mL HyA. The results from the in vitro experiment were used as culture parameters for the in vivo analysis. Composite 5% HyA/collagen I or collagen I-only hydrogels were used to seed chondrocytes into SFF-fabricated scaffolds made of PPF with designed cubic or ellipsoidal pore geometry. The scaffolds were implanted subcutaneously in immunocompromised mice for 4 weeks. Histomorphometric analyses of sections stained with Safranin O were used to quantify the amount of ECM deposited by cells in the scaffolds. Scaffolds seeded with 5% HyA/collagen hydrogels had significantly greater areas of positive Safranin O staining (approximately 60%, compared with 30% for scaffolds with collagen I hydrogels only), indicating that greater numbers of chondrocytes retained their metabolic activity in the ectopic environment. These scaffolds also had greater stain intensities (corresponding to greater amounts of sGAG in the ECM) than their counterparts seeded with collagen I hydrogels alone. Significant differences in matrix production were not found between the scaffold pore designs. Overall, these results indicate that a combination of composite HyA hydrogels and designed SFF scaffolds could provide a functional tissue-engineered construct for cartilage repair with enhanced tissue regeneration in a load-bearing scaffold.

Chondrogenic differentiation of human bone marrow-derived mesenchymal stem cells in a simulated osteochondral environment is hydrogel dependent

“…Still chondrogenesis took place in alginate/HA, indicated by comparable levels of Col2 and ACAN gene expression after 28 d of culture in chondrogenic medium for all four hydrogels. Since positive findings were reported for the use of HA both clinically and in laboratory settings (Grigolo et al, 2001;Kavalkovich et al, 2002;Facchini et al, 2006;Liao et al, 2007;Rampichova et al, 2010;Kim et al, 2011;Erickson et al, 2012), we still believe that HA can be beneficial for cartilage tissue engineering purposes, albeit not in the current combination of high molecular weight HA with alginate. Therefore, besides alginate and fibrin, HA-pNIPAM was used to prepare hBMSC-hydrogel constructs in the osteochondral biopsy model.…”

Hydrogels for the Repair of Articular Cartilage Defects

“…72 For example, the addition of hyaluronan to collagen I hydrogels increased markers of cartilage formation by encapsulated chondrocytes in a subcutaneous implantation model. 73 The addition of hyaluronan to cell culture medium increased deposition of type II collagen and glycosaminoglycans, markers of the cartilage phenotype, by chondrocytes encapsulated in alginate hydrogels, 82 but decreased such markers by chondrocytes encapsulated in collagen hydrogels. 83 Other studies have shown that the addition of hyaluronan to alginate hydrogels caused an increase in the expression of type I collagen, a marker of fibrous scar-like cartilage formation.…”

Anti-Inflammatory Strategies in Cartilage Repair

“…Of the synthetic polymers, those derived from poly(a-hydroxy esters), 136 poly(propylene fumarates), 137 polyurethanes, 138 PGA, 139 and poly(lactideco-glycolide) 140 have been used for cartilage regeneration; however, inflammatory reaction due to synthetic scaffolds that impaired the quality of regenerated cartilage has also been reported in some studies. 141,142 Natural matrix scaffolds Naturally occurring polymers as scaffolds offer options for cartilage tissue engineering due to biocompatibility, biodegradability, low toxicity of degradation by-products, and plasticity in processing into a variety of material formats.…”