Dynamic compression stimulates proteoglycan synthesis by mesenchymal stem cells in the absence of chondrogenic cytokines

Abstract

The objective of this study was to evaluate the effect of dynamic compression on mesenchymal stem cell (MSC) chondrogenesis. Dynamic compression was applied to agarose hydrogels seeded with bone marrow-derived adult equine MSCs. In the absence of the chondrogenic cytokine transforming growth factor beta (TGFbeta), dynamic compression applied for 12 h per day led to significantly greater proteoglycan synthesis than in unloaded TGFbeta-free cultures, although at a rate that was approximately 20% to 35% of unloaded TGFbeta cultures. These data suggest that the emergence of aggrecan dominated a chondrogenic response to loading as increases in proteoglycan synthesis. Cross-sectional analyses were conducted to subjectively identify potential spatial distributions of heterogeneous differentiation. In loaded samples, cell viability and metachromatic staining was low near the porous compression platen interface but increased with depth, reaching levels in the lower portion of the hydrogel that resembled unloaded TGFbeta cultures. These results suggest that the combination of high hydrostatic pressure and low dynamic strain and fluid flow had a stronger effect on chondrogenesis than did low hydrostatic pressure coupled with high dynamic strain and fluid flow. Next, the 12-h per day loading protocol was applied in the presence of TGFbeta. Biosynthesis in loaded cultures was less than in unloaded TGFbeta samples. Taken together, these data suggest that the duration of loading necessary to stimulate mechanoinduction of MSCs may not be optimal for neo-tissue accumulation in the presence of chondrogenic cytokines.

FIG. 1.

Extracellular matrix synthesis in response to dynamic compression in the absence of transforming growth factor beta (TGFβ) after 21 days of culture. Data for each loading protocol were calculated from cultures established from three donor animals. For each assay, significant differences between conditions were denoted by the labels ‘a,’ ‘b,’ and ‘c.’

FIG. 2.

Viable cell distribution and proteoglycan deposition over full-thickness (3 mm) sections of samples loaded using the 12-h/d protocol in the absence of TGFβ (Day 21). Insets show representative sections from unloaded TGFβ− and TGFβ+ controls. Color images available online at www.liebertonline.com/ten.

FIG. 3.

Viable cell distribution and proteoglycan deposition adjacent to the impermeable base after 21 days of the 12-h/d loading protocol in the absence of TGFβ. Color images available online at www.liebertonline.com/ten.

FIG. 4.

Extracellular matrix synthesis in response to dynamic compression in the presence of TGFβ after 15 days of culture. Dynamic compression samples were loaded using the 12-h/day protocol. Data were calculated from cultures established from three donor animals. For each assay, significant differences among conditions were denoted by the labels ‘a’ and ‘b.’

FIG. 5.

Proteoglycan deposition over full-thickness (3 mm) sections of samples maintained in medium containing TGFβ (Day 15). Dynamic compression samples were loaded using the 12-h/d protocol. Color images available online at www.liebertonline.com/ten.