gms | German Medical Science

Objectives: In osteoarthritis (OA), pro-inflammatory conditions are widely acknowledged to play a crucial role in the progression of the disease. Also when aiming at cell-based therapy to fill cartilage defects in OA, possible effects of inflammatory mediators such as IL-1β on implanted tissue-engineered (TE) constructs have to be taken into account. Recent studies have shown the polyphenolic phytoalexin resveratrol (RSV) to have potent anti-inflammatory properties and revealed molecular signaling pathways involved. However, long-term effects on TE constructs remain largely unexplored. Therefore, in this study the effects of RSV on 3D pellet constructs made from articular chondrocytes were investigated in the presence of IL-1β.

Methods: Porcine articular chondrocytes were isolated from the femorotibial joint, passaged, and pellets with 2x10⁵ cells were formed. Pellet constructs were then cultured in a serum-free medium and stimulated either with IL-1β (1 - 10 ng/ml) or RSV (50 μM) alone, or were co-treated with both agents. Untreated samples served as control (Ctrl). In one set of experiments, pellets immediately received treatment for 14 days, whereas in other experiments, pellets were pre-cultered in serum-free medium for 7 days followed by culture with treatments for 14 days. All constructs were analyzed using histology, immunohistochemistry, and quantitative biochemical assays.

Results and conclusion: In experiments without pre-culture, IL-1β (10 ng/ml), as compared to controls, drastically decreased both pellet size and glycosaminoglycan (GAG) content as normalized to DNA (Ctrl: 92.04±8.94 μg/μg, IL-1β: 8.35±0.96 μg/μg GAG/DNA; p<0.01). Co-treatment of IL-1β-stimulated constructs with RSV significantly counteracted the pro-inflammatory catabolism and led to partial rescue of GAG content (43.44±7.20 μg/μg GAG/DNA; p<0.01). Pellets undergoing 7 days of pre-culture followed by 14 days of treatment showed a similarly strong decrease of GAG content when treated with IL-1β (Ctrl: 89.38±7.14 μg/μg, IL-1β: 12.24±1.06 μg/μg GAG/DNA; p<0.01), but an even more pronounced rescue by co-treatment of IL-1β-stimulated constructs with RSV up to control levels (99.31±13.59 μg/μg GAG/DNA; p<0.01).

Immunohistochemistry showed abundant collagen II content in all samples. Undesired collagen I and degradative enzyme MMP-13 were strongly increased in IL-1β-treated samples. However, strikingly, addition of RSV completely reversed these effects. Collagen X was detected in pellets treated with RSV only, but in no other group including pellets co-treated with IL-1β and RSV.

Our results indicate that the chondroprotective effects of RSV counteract IL-1β-mediated degradation of 3D chondrocyte constructs in a long-term culture. Further investigations in vitro as well as evaluation in an in vivo model including inflammatory conditions appear desirable. RSV may be used as a potential co-treatment when implanting tissue-engineered constructs in an inflammatory environment such as in OA.