Mitochondrial membrane potential and reactive oxygen species content of endothelial and smooth muscle cells cultured on poly(ε-caprolactone) films

A transitory but significant stimulation of mitochondrial activity, increase of reactive oxygen species (ROS) and oxidative stress were previously observed in L929 fibroblasts cultured on poly(ε-caprolactone) (PCL) films. ROS, mainly formed in mitochondria, play a physiological role but an excessive production can promote endothelial dysfunction, cause oxidative injury to vascular cells, oxidize lipoproteins and accelerate atherothrombogenesis. On the other hand, mitochondria have a crucial position in programmed cell death control and are responsible for ATP synthesis through the coupling of oxidative phosphorylation to respiration. This coupling requires the existence of a mitochondrial membrane potential (Δψm). The aim of the present study was to evaluate by flow cytometry the ROS content and Δψm of both endothelial (EC) and smooth muscle cells (SMC) cultured on PCL films as a potential substrate for vascular graft development. Cell size, internal complexity and cell cycle were also analyzed to detect the possible appearance of the subG1 cell fraction, characteristic of apoptotic cells. The effect of treating PCL films with NaOH before culture was also studied. PCL decreases the ROS content of EC during the culture but produces an increase of these levels in SMC after 7 days. PCL also induces variations of Δψm which show a significant parallelism with the changes observed in ROS levels proving the importance and sensitivity of these measurements as indicators of the mitochondrial function. The treatment of PCL with NaOH decreases these effects demonstrating the benefits of increasing the surface hydrophilicity before cell culture which improves cell adhesion and proliferation and reduces oxidative stress. Since no important changes have been detected in subG1 fraction of EC and SMC cultured on either PCL or PCL–NaOH, the changes of Δψm observed in the present study cannot be related to apoptosis. These results confirm the potential utility of PCL as a suitable scaffold in Vascular Tissue Engineering.