Wang Xing-Hua
Department of Ultrasound, Second Hospital of Shanxi Medical University, Taiyuan, China.
J Ultrasound Med. 2011 Mar;30(3):325-32. doi: 10.7863/jum.2011.30.3.325.
The mechanism by which Optison (an albumin-shelled, octafluoropropane gas-filled microbubble contrast agent; Amersham Health, Amersham, England) enhances gene transfection in skeletal muscle in vivo with or without ultrasound (US) is unclear. The possible mechanisms were investigated by experimenting with different constituents, both with and without US.
Plasmid DNA (10 μg) encoding green fluorescent protein was mixed with Optison or its constituents dissolved in saline (in an equivalent concentration as in Optison) and injected into the tibialis anterior muscle of mice with or without adjunct US (1 MHz, 2 W/cm², 30 seconds, and 20% duty cycle). The efficiencies of green fluorescent protein transgene expression were determined under different experimental conditions: (1) plasmid plus saline as a negative control; (2) plasmid plus Optison as a positive control; (3) plasmid plus heat-treated Optison (without microbubbles); (4) plasmid plus human serum albumin; (5) plasmid plus N-acetyltryptophan; and (6) plasmid plus caprylic acid. Transfection efficiency was assessed by counting the maximum number of green fluorescent protein-positive fibers. Tissue damage was assessed by measuring the damaged area on serial sections.
Heat-treated Optison with or without US and albumin with US showed similarly high levels of transgene expression as Optison in mouse muscle without substantially increased tissue damage. N-Acetyltryptophan and caprylic acid had no effect on the delivery of plasmid green fluorescent protein into mouse muscle but instead showed the potential to increase tissue damage.
These data suggest that US and albumin separately potentiate transfection in this model. The combination of albumin and perfluoropropane is highly effective, which probably explains why Optison is so effective.
尚不清楚Optison（一种白蛋白包裹、填充八氟丙烷气体的微泡造影剂；安进公司，英国阿默舍姆）在有或无超声（US）的情况下增强体内骨骼肌基因转染的机制。通过对不同成分进行有无超声的实验来研究可能的机制。
将编码绿色荧光蛋白的质粒DNA（10μg）与溶解于生理盐水的Optison或其成分（浓度与Optison中的等效）混合，在有或无辅助超声（1MHz，2W/cm²，30秒，占空比20%）的情况下注入小鼠胫前肌。在不同实验条件下测定绿色荧光蛋白转基因表达效率：（1）质粒加生理盐水作为阴性对照；（2）质粒加Optison作为阳性对照；（3）质粒加热处理的Optison（无微泡）；（4）质粒加人血清白蛋白；（5）质粒加N - 乙酰色氨酸；（6）质粒加辛酸。通过计数绿色荧光蛋白阳性纤维的最大数量评估转染效率。通过测量连续切片上的损伤面积评估组织损伤。
有或无超声的热处理Optison以及有超声的白蛋白在小鼠肌肉中显示出与Optison相似的高转基因表达水平，且组织损伤未显著增加。N - 乙酰色氨酸和辛酸对质粒绿色荧光蛋白向小鼠肌肉的递送无影响，但显示出增加组织损伤的可能性。
这些数据表明在该模型中超声和白蛋白分别增强转染。白蛋白和全氟丙烷的组合非常有效，这可能解释了Optison为何如此有效。
