The role of the extracellular matrix and mechanical strain on gene expression of engineered smooth muscle tissue

The field of tissue engineering holds great promise for the treatment of cardiovascular disease. Cardiovascular tissues reside in a mechanically dynamic environment and it is therefore important to understand how vascular cells respond to mechanical signals from the extracellular matrix (ECM) to which they are adhered. The goal of this thesis is to provide a better understanding of the effects of mechanical strain on cardiovascular cell gene expression, particularly in terms of genes associated with the calcification process in the vasculature. Tissues were engineered by seeding smooth muscle cells (SMCs) into three-dimensional scaffolds. Mechanical strain was applied using a custom-made device to exert a uniaxial strain of 7% at a frequency of 1 Hz. SMCs utilized distinct integrin receptors to adhere to various substrates and cellular gene expression was regulated by the scaffold chemistry. These cells were found to express a variety of bone-associated genes, such as osteopontin, matrix gla protein (MGP), alkaline phosphatase, and the transcription factor CBFA-1. Strikingly, however, expression of these genes was downregulated in tissues exposed to cyclic strain at all time points ranging from 5 to 150 days. Further, long-term strain played a protective role against calcification, as unstrained tissues exhibited increased calcium deposition compared to strained tissues. The mechanism of strain-induced osteopontin and MGP gene expression by SMCs was also studied by investigating the involvement of the ECM and matrix-associated protein kinase (MAPK) signaling pathways. The MAPK signal transduction pathways ERK and p38 were studied and neither pathway regulated the expression of either osteopontin or MGP with strain application. However, blocking ERK signaling inhibited the expression of MGP, implicating the involvement of that pathway in the regulation of MGP expression. Overall, the results in this thesis suggest that without an appropriate mechanical environment, SMCs in three-dimensional culture undergo a phenotypic conversion to an osteoblast-like pattern of gene expression. These studies shed light onto how mechanical strain is transduced through the cell and regulates the gene expression of known mediators in the vascular calcification process. Understanding the role of strain in the vasculature can provide insight into causes of disease and lead to better treatment options.