Study of cell death induced by cell membrane localized sonodynamic therapy

Sonodynamic therapy (SDT), based on synergistic effect of low intensity ultrasound and sonosensitizer, is a potential noninvasive approach for the treatment of cancers. It is shown that the subcellular distribution of sonosensitizer are varied with their properties and incubation time. However, the responses of cancer cells to localized SDT is still unclear. The aim of this study is to reveal bioeffect of cell membrane localized SDT. Before ultrasound treatment, Homo sapiens epithelial cells, HeLa were incubated 1 hour with protoporphyrin IX (PpIX, accumulated on cell membrane in 1 hour). The subcellular distribution of PpIX was visualized after incubating of 1 hour by confocal microscopy. Then cells were treated by low intensity ultrasound (frequency of 1.0MHz, Ispta of 500mW/cm², and duty circle of 20%) for 10 minutes. After SDT treatment, FITC Annexin V/PI stain and flow cytometry analysis were employed for the analysis of the necrosis and apoptosis ratio induced by cell membrane localized SDT. Confocal images showed that PpIX adhered to membrane after incubation of 1 hour. Flow cytometry results demonstrated that there were 41.8% necrosis and 5.12% apoptosis when loaded with PpIX after SDT treatment. These results indicated that membrane localized SDT tends to induce necrosis and provide us an alternative protocol for SDT.