叶酸靶向液态氟碳脂质纳米粒的制备及其超声分子显像卵巢癌实验

Preparation of folic acid targeted liquid fluorocarbon lipid nanoparticles and application in ultrasound molecular imaging of ovarian cancer

目的制备叶酸修饰的液态氟碳脂质纳米粒,并探讨其在不同温度下的稳定性及在体内外模型中靶向结合卵巢癌SKOV3细胞的效能。方法应用薄膜水化法和旋转蒸发法制备叶酸靶向液态氟碳脂质纳米粒。观察纳米粒的基本特性及在4℃和37℃水浴条件下的稳定性。分为靶向组(叶酸靶向)、非靶向组(普通磷脂)及拮抗组(游离叶酸干预),通过卵巢癌SKOV3细胞,进行脂质纳米粒的体外靶向实验。并将8只荷SKOV3肿瘤裸鼠分为A组(靶向)和B组(非靶向),进行脂质纳米粒的体内靶向实验。结果本实验制备的纳米粒大小均匀,粒径约(321.20±67.21)nm,表面电位-50mV。在4℃条件下,纳米粒可稳定保存48h;在37℃水浴条件下,1h内纳米粒的粒径无明显改变。靶向组纳米粒与SKOV3细胞的结合率最高,拮抗组最低。靶向组肿瘤细胞内可见大量被荧光染料DiI标记为红色的纳米粒;非靶向组和拮抗组仅见少量红色荧光。A组裸鼠肿瘤局部可见较多被荧光染料DiR标记为红色的纳米粒,而B组肿瘤局部未见明显红色荧光。且A组裸鼠各重要脏器离体荧光成像肿瘤荧光明显强于B组。结论本实验制备的叶酸靶向液态氟碳脂质纳米粒具有一定的稳定性,能够靶向结合于卵巢癌SKOV3细胞。

Objective To prepare perfluoropentane nanodroplets modified by folate and encapsulated by lipid membrane （FA-NDs）, and to investigate the nanodroplets stability in different temperature and its target performance of SKOV3 tumor cells in vitro and in vivo. Methods FA-NDs nanodroplets were fabricated with lipid film hydration and rotary evapo- ration methods. The nanodroplets stability was evaluated at 4℃ and 37℃, respectively. In vitro target test was demonstra- ted with co-incubation of SKOV3 tumor cells and nanodroplets which were divided into targeted group, non-targeted group and antagonism group. Totally 8 tumor-bearing mice were divided into group A （targeted） and group B （non-targeted） for in vivo target experiment of nanodroplets. Results The nanodroplets were successfully prepared with good size uniformity. The mean diameter of prepared nanodroplets was （321.20 ：t： 67.21）nm. And the average surface zeta potential was --50 inV. The nanoparticles remained stable for 48 h at 4℃ and 1 h at 37℃. In vitro targeted experiments showed the highest binding rate for SKOV3 cells existed in targeted group and the lowest binding rate existed in antagonism group. In targeted group, numerous red dots representing FA-NDs dyed with DiI were visible around the cell membrane and in the cytoplasm of SKOV3 cells. While minimal red signal was observed in non-targeted and antagonized groups. The numerous red fluorescence representing FA-NDs dyed with DiR were visible in mice tumor of group A, but there was nearly no red fluorescence in non-targeted group. Ex vivo fluorescence intensity of mice organs and tumors in the targeted group was sig- nificantly stronger than that in non-targeted group. Conclusion FA-NDs nanodroplets prepared in this study are stable and can be used in targeting SKOV3 tumor cells specifically.