Damage effects of protoporphyrin IX – Sonodynamic therapy on the cytoskeletal F-actin of Ehrlich ascites carcinoma cells

In this study, we report evidence of the damage effects of sonodynamic therapy (SDT) on a novel intracellular target, cytoskeletal F-actin, that has great importance for cancer treatment. Ehrlich ascites carcinoma (EAC) cells suspended in PBS were exposed to ultrasound at 1.34 MHz for up to 60 s in the presence and absence of protoporphyrin IX (PPIX). To evaluate the polymeric state and distribution of actin filaments (AF) we employed FITC-Phalloidin staining. The percentage of cells with intact AF was decreased with 10–80 μΜ PPIX after ultrasonic exposure, while only few cells with disturbed F-actin were observed with 80 μΜ PPIX alone. The fluorescence intensity of FITC-Phalloidin labeled cells was detected by flow cytometry. The morphological changes of EAC cells were observed by scanning electron microscope (SEM). The nuclei were stained with Hoechst 33258 to determine apoptosis. Cytoskeletal F-actin and cell morphological changes were dependent on the time after SDT. Some cells suffered deformations of plasma membrane as blebs that reacted positively to FITC-Phalloidin at 2 h after SDT treatment. Many of the cells showed the typically apoptotic chromatin fragmentation. The alterations were more significant 4 h later. Our results showed that cytoskeletal F-actin might represent an important target for the SDT treatment and the observed effect on F-actin and the subsequent bleb formation mainly due to apoptosis formation due to the treatment.