Regulating orientation and phenotype of primary vascular smooth muscle cells by biodegradable films patterned with arrays of microchannels and discontinuous microwalls

Vascular smooth muscle cells (vSMCs) cultured in vitro are known to exhibit phenotype hyperplasticity. This plasticity is potentially very useful in tissue engineering of blood vessels. The synthetic phenotype is necessary for cell proliferation on the tissue scaffold but the cells must ultimately assume a quiescent, contractile phenotype for normal vascular function. In vitro control of vSMC phenotype has been challenging. This study shows that microchannel scaffolds with discontinuous walls can support primary vSMC proliferation and, when the cells reach confluence inside the channels, transform the cell phenotype towards greater contractility and promote cell alignment. A thorough time-resolved study was undertaken to characterize the expression of the contractile proteins alpha-actin, calponin, myosin heavy chain (MHC) and smoothelin as a function of time and initial cell density on microchannel scaffolds. The results consistently indicate that primary vSMCs cultured on the microchannel substrate substantially align parallel to the microwalls, become more elongated and significantly increase their expression of contractile proteins only when the cells reach confluence. MHC immunostaining was visible in the micropatterned cells after confluence but not in flat substrate cells or non-confluent micropatterned cells, which further verifies the increased contractility of the confluent channel-constrained vSMCs. The higher total amount of deposited elastin and collagen in confluent flat cultures than in confluent micropatterned cultures also provides confirmation of the higher contractility of the channel-constrained cells. These results establish that our microchanneled film can trigger the switch of primary vSMCs from a proliferative state to a more contractile phenotype at confluence.