低频超声联合微泡造影剂诱导人前列腺癌PC3细胞及DU145细胞自噬的实验研究

Autophagy induced by low-intensity ultrasound combined with microbubbles in human prostate cancer cells PC3 cells and DU145 cells

目的研究低频超声联合微泡造影剂对人雄激素非依赖性前列腺癌PC3细胞及DU145细胞自噬的影响。方法 2种细胞均各分为空白对照组、单纯微泡组、单纯低频超声组和低频超声联合微泡组4组,每组设3个复孔。单纯微泡组加200μl的微泡造影剂混匀,不进行超声辐照;单纯低频超声组以声功率80 mW的20 kHz低频超声,不加造影剂,连续波辐照60 s;低频超声联合微泡组加200μl的微泡造影剂,混匀后以声功率80 mW的20 kHz低频超声,连续波辐照60 s;空白对照组不进行任何处理,辐照后的细胞继续培养24 h,吖啶橙染色荧光显微镜与透射电镜观察细胞自噬泡。结果经吖啶橙染色后,荧光显微镜下,2种细胞低频超声联合微泡组的胞质内可见大量红色荧光的酸性囊泡,明显多于其余3组;单纯低频超声组红色荧光的酸性囊泡量亦多于空白对照组,空白对照组与单纯微泡组红色荧光的酸性囊泡量差异不大。透射电镜下,2种细胞的低频超声联合微泡组细胞细胞核基本正常,但胞质内出现大量由双层膜包裹的自噬泡或自噬体,而空白对照组内的细胞形态基本正常,胞质内未见到明显的自噬泡的形成。结论低频超声联合微泡造影剂能明显诱导人雄激素非依赖型前列腺癌PC3细胞及DU145细胞的自噬。

Objective To investigate the effect of 20 kHz low-intensity ultrasound combined with microbubbles on autophagy of both PC3 cells and DU145 cells. Methods Ultrasound with a frequency of 20 kHz and intensity of 80 mW in continuous wave mode was used. Both PC3 cells and DU145 cells were divided into four groups, including control group （A）, microbubbles group （B）, ultrasound group （C） and ultrasound combined with microbubbles group （D）. Twenty-four hours after treatment, the acid vesicular organelles were detected by acridine orange lfuorescence staining, and transmission electron microscopy （TEM） was used to observe autophagosomes. Results Acridine orange lfuorescence staining showed formation of acid vesicular organelles （AVOs） in the cytoplasm with normal nucleus in both PC3 cells and DU145 cells in group D, while in group A cells were basically normal. Lots of autophagosomes with double-membrane structure were detected by transmission electron microscope in group D. Conclusions Low-frequency and low-intensity ultrasound combined with microbubbles can induced autophagy in both PC3 cells and DU145 cells.