Low-intensity ultrasound-induced cellular destruction and autophagy of nasopharyngeal carcinoma cells

Abstract

Abstract. Ultrasound therapy, as a non-invasive modality, has been attracting extensive attention in the management of malignant tumors. The present study aimed to investigate low-intensity ultrasound-induced cellular destruction and autophagy in nasopharyngeal carcinoma cells in vitro. Nasopharyngeal carcinoma CNE2 cells were subjected to ultrasound exposure, as tumor model cells, at an intensity of 1.35 W/cm 2 . Cytotoxicity was investigated 24 h after ultrasound treatment. Nuclear damage was observed using nuclear staining with Hoechst 33258. Mitochondrial dysfunction was measured using confocal laser scanning microscopy with rhodamine123 staining. Mitochondrial morphology and autophagy were observed using transmission electron microscopy (TEM). Low-intensity ultrasound significantly killed CNE2 cells proportional to the ultrasonic treatment time. Upon nuclear staining, nuclear condensation and typical apoptotic bodies were noted in the CNE2 cells exposed to ultrasound wave for 12 sec. A collapse in mitochondrial membrane potential was noted in the treated cells. Upon TEM, swollen mitochondria, more vacuoles and autophagy were noted after ultrasound treatment. Our findings demonstrate that low-intensity ultrasound significantly damages CNE2 cells and emphasize that autophagy may be an important event in ultrasound-induced cell death. IntroductionNasopharyngeal carcinoma (NPC) is one of the most common malignant tumors in Southeast Asia, including Hong Kong. Current treatments for NPC, such as surgery, radiotherapy and chemotherapy, have serious side effects with limited success (1-4). Fortunately, continued research efforts have spawned novel therapies.With the development of transducer design and measurement technology, ultrasound as a safe alternative and non-invasive therapeutic approach has been widely applied in medicine (5). High-intensity focal ultrasound (HIFU) produces a high temperature that directly or indirectly deactivates tumor cells/tissues and is therefore undergoing development as a noninvasive therapeutic modality in the management of cancer (6). Recently, the applications of low-intensity ultrasound in tumors and vascular diseases have been widely investigated. Emerging evidence has confirmed that low-intensity ultrasound markedly inhibits the proliferation and clone formation of tumor cells through heat, mechanical effects and acoustic cavitation (7,8). Fitzgerald et al demonstrated that therapeutic ultrasound markedly decelerated cellular proliferation and decreased in-stent hyperplasia (9). Takeuchi et al reported that ultrasound energy inhibits the proliferation of mouse T lymphoma cells at an intensity of 350 mW/cm 2 and human histiocytic lymphoma cells (U-937) at an intensity of 700 mW/ cm 2 . The morphological changes included apoptosis, such as the shrinking of cells and formation of apoptotic bodies (10). However, it remains unclear which types of mechanisms are involved in these bioeffects. Recent studies have shown that autophagy is involved in the pathogenesis of tumors. The ind...

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