The role of mechanical stimuli in engineered smooth muscle tissue development.

Abstract

Engineering new tissues (e.g., blood vessels) may be possible by transplanting cells on three-dimensional, biodegradable scaffolds. It will be critical to engineer functional smooth muscle (SM) tissue if one hopes to successfully replace the large number of tissues containing a SM component (e.g., blood vessels, intestines, and bladder) with engineered equivalents. This thesis was based on the hypothesis that cyclic mechanical strain regulates gene expression of smooth muscle cells (SMCs) and the formation, structure, and function of engineered SM tissue. A model system was developed to test this hypothesis by optimizing cell seeding and culture methods on three-dimensional scaffolds, and selecting appropriate cyclic strain conditions and elastic three-dimensional scaffolds for this system. SM tissues with biochemical compositions similar to native SM could be engineered by optimizing cell seeding and culture methods. However, the structure of the tissues developed in mechanically static conditions and did not resemble native SM. To address the hypothesis guiding this thesis, SM tissues were engineered with type I collagen sponges and bonded PGA scaffolds, and subjected to cyclic strain with an amplitude of 7% and a frequency of 1Hz for up to 20 weeks. SMCs' response to cyclic strain strongly dependent on the interaction with extracellular matrix (ECM) molecules utilized for the cell adhesion. Cyclic strain significantly increased cell proliferation and ECM (elastin and collagen) synthesis on fibronectin-coated, bonded PGA scaffolds, but not on type I collagen sponges in serum-free medium. In chronic mechanical stimuli, cyclic strain stimulated the process of engineered SM tissue formation by increasing cell proliferation (up to 17 +/- 4%) and the synthesis rates of elastin (up to 92 +/- 10%) and collagen (up to 90 +/- 31%) compared to control (subjected to no cyclic strain) on both bonded PGA scaffolds and type I collagen sponges in serum-containing medium. Cyclic mechanical strain also stimulated the organization in engineered tissues by inducing orientation of SMCs. Importantly, the direction of SMC alignment in engineered tissues was regulated by the amplitude of cyclic strain and the duration of cyclic strain application. Cyclic strain with the amplitude reduced from 7% to 3.5% for 10 weeks decreased cell alignment angle from 53 +/- 5° to 0° (parallel) to cyclic strain direction, while no alignment was observed in the control tissues. Cyclic strain with an amplitude of 7% for a longer period (20 weeks) decreased the cell alignment angle from 53 +/- 5° on week 10 to 20 +/- 3° to cyclic strain direction. In addition, cyclic strain dramatically enhanced the mechanical properties of engineered SM tissues compared to control. The ultimate tensile strength and Young's modulus of the engineered SM tissues subjected to cyclic strain for 20 weeks was 12 +/- 3-fold and 34 +/- 9-fold higher, respectively, than those of the control tissues. This thesis demonstrated that cyclic mechanical strain plays a critical role in the development of engineered SM tissue with correct structure and function. This thesis suggests that functional SM tissue could be engineered by application of appropriate mechanical signals.