Vinculin is a key player in sensing and responding to external mechanical cues such as extracellular matrix stiffness. Increased matrix stiffness is often associated with certain pathological conditions including hypertension induced cellular cytoskeleton changes in vascular smooth muscle (VSM) cells. However, little is known on how stiffness affects cytoskeletal remodeling via vinculin in VSM cells. Thus, we utilized matrices with elastic moduli that simulate vascular stiffness in different stages of hypertension to investigate how matrix stiffness regulates cell cytoskeleton via vinculin in synthetic VSM cells. Through selecting a suitable reference gene, we found that an increase in physiologically relevant extracellular matrix stiffness (2-50 kPa) downregulates vinculin gene expression but upregulates vinculin protein expression. This discrepancy, which was not observed previously for non-muscle cells, suggests that the vinculin-mediated mecahnotransduction mechanism in synthetic VSM cells may be more complex than those proposed for non-muscle cells. Also adding to previous findings, we found that VSM cell growth may be impeded by substrates that are either too soft or too rigid. (C) 2018 Elsevier Inc. All rights reserved.