Honda Hidemi, Zhao Qing-Li, Kondo Takashi
Department of Radiological Sciences, Faculty of Medicine, Toyama Medical and Pharmaceutical University, Toyama, Japan.
Ultrasound Med Biol. 2002 May;28(5):673-82. doi: 10.1016/s0301-5629(02)00509-4.
Human histiocytic lymphoma U937 cells were exposed to continuous 1-MHz ultrasound (US) for therapeutic use, (0 approximately 6.5 W/cm(2) (I(SPTA)). Apoptosis and its related end points were examined by flow cytometry. Fraction of cells with low mitochondria membrane potential were observed after sonication and significant superoxide and peroxide formation, increased activity of caspase-3, and DNA fragmentation revealed biochemically, were also found. The fraction of early apoptosis and secondary necrosis increased with the incubation time after sonication. Early apoptosis observed at 6 h after sonication reached its maximum at 2 min of sonication and gradually decreased. On the other hand, secondary necrosis increased with the duration of sonication. When the effects of dissolved gases, Ar, N(2), O(2), air, N(2)O and CO(2), on free radical formation due to inertial cavitation were investigated by electron spin resonance (ESR) spin trapping, formation of hydroxyl radicals and hydrogen atoms was found in solutions saturated with Ar, N(2), O(2) and air, but not with N(2)O and CO(2). Apoptosis induced by US was also dependent on the dissolved gases in the order Ar = N(2) = O(2) = air >> N(2)O = CO(2) approximately 0. These results suggest that US-induced apoptosis, which is mitochondria-caspase dependent, was linked to inertial cavitation. However, quantities of free radicals did not influence the fraction of early apoptosis and secondary necrosis. When the cells were sonicated in the presence of an echo contrast agent, Levovist; synergistic enhancement of secondary necrosis induced by US was observed at concentrations of more than 20 mg/mL. In contrast, an additive increase of early apoptosis was observed in the combined treatments. These results suggest that Levovist; acting as cavitation nuclei, enhances secondary necrosis induced by US due to an increase in the membrane damage.
将人类组织细胞淋巴瘤U937细胞暴露于连续1兆赫兹的超声（US）下用于治疗，强度为0至约6.5瓦/平方厘米（I(SPTA)）。通过流式细胞术检测细胞凋亡及其相关终点。超声处理后观察到线粒体膜电位低的细胞比例，同时还发现有显著的超氧化物和过氧化物形成，生化检测显示半胱天冬酶-3活性增加以及DNA片段化。早期凋亡和继发性坏死的比例随超声处理后的孵育时间增加。超声处理后6小时观察到的早期凋亡在超声处理2分钟时达到最大值，随后逐渐下降。另一方面，继发性坏死随超声处理时间延长而增加。当通过电子自旋共振（ESR）自旋捕获研究溶解气体氩气（Ar）、氮气（N₂）、氧气（O₂）、空气、一氧化二氮（N₂O）和二氧化碳（CO₂）对惯性空化引起的自由基形成的影响时，发现氩气、氮气、氧气和空气饱和的溶液中会形成羟基自由基和氢原子，而一氧化二氮和二氧化碳饱和的溶液中则不会。超声诱导的凋亡也取决于溶解气体，顺序为氩气 = 氮气 = 氧气 = 空气 >> 一氧化二氮 = 二氧化碳≈0。这些结果表明，超声诱导的依赖线粒体 - 半胱天冬酶的凋亡与惯性空化有关。然而，自由基的量并不影响早期凋亡和继发性坏死的比例。当在回声造影剂声诺维（Levovist）存在下对细胞进行超声处理时，在浓度超过20毫克/毫升时观察到超声诱导的继发性坏死有协同增强作用。相反，联合处理中观察到早期凋亡呈相加增加。这些结果表明，声诺维作为空化核，由于膜损伤增加而增强了超声诱导的继发性坏死。
