Influence of pressurized cyclic stretch and endothelial cell presence on multipotent stem cell osteogenic commitment

Abstract

Applied mechanical stretch and blood vessel invasion are key stimuli to which progenitor cells are exposed in post-natal endochondral bone formation. Understanding the combined effects of cyclic stretch and endothelial cell (EC) presence on multipotent stem cell (MSC) osteogenesis therefore has the potential to lead to improved MSC-based bone regeneration strategies. Toward this goal, 10T1/2 mouse MSCs were encapsulated in tubular poly(ethylene glycol) diacrylate [PEGDA] hydrogels with moduli within the “osteogenic” range in order to induce osteogenesis. Half of the constructs were fabricated with a luminal EC layer. All of the EC⁺ (EC⁺/dyn⁺) and half of the EC⁻ constructs (EC⁻/dyn⁺) were subjected to pressurized cyclic stretch in the absence of osteogenic media supplements, with remaining EC⁻ constructs (EC⁻/dyn⁻) serving as static controls. At day 10 of culture, expression of the bone extracellular matrix protein osteopontin was over 3.3- and 1.9-fold higher in the EC⁺/dyn⁺ and EC⁻/dyn⁺ constructs, respectively, relative to day 0. At day 22 of culture, osteopontin levels could not be statistically distinguished from day 0 in the EC⁺/dyn⁺ constructs and were one-third less than day 0 in the EC⁻/dyn⁺ constructs. In contrast, at day 22 levels of an osteogenic marker alkaline phosphatase (AP) were over 2.4- and 1.4-fold higher in the EC⁺/dyn⁺ and EC⁻/dyn⁺ constructs, respectively, relative to day 0. Furthermore, at day 22 matrix mineralization in both dynamic groups was increased over 2.5-fold and over 9-fold relative to the EC⁻/dyn⁻ and day 0 groups, respectively. Cumulatively, these results suggest that pressurized cyclic stretch alone significantly increases the rate/degree of osteogenesis relative to static culture. However, EC presence combined with pressured cyclic stretch appears to further enhance the rate/degree of MSC osteogenesis and/or to support a distinct osteogenic “fingerprint” compared to that promoted by cyclic stretch alone.