普鲁兰基纳米药物载体透过胎盘屏障的机制及其调控策略研究

It is a big challenge that how to reduce the possible embryo toxicity because of the drug crossing the placental barrier during pregnancy. It is very necessary and urgent to study the mechanism and controlling strategy of crossing the placental barrier for nano drug carrier used in perinatal medicine field. Pullulan acetate nanoparticles (PANs) are safe and showed drug slow releasing property. On the previous work, we build the placental choriocarcinoma (BeWo) placental barrier model, observe the ability and mechanism of crossing the placental barrier of PANs on the BeWo model. Fluorescence microscope, transmission electron microscope and laser confocal method were used to observe the transport characterization mechanism. Detect the level of cytokine by ELISA, then elucidate the correlation between the transport rate of PANs, endocytosis inhibitors and the level of related cytokine. Discuss the possibility of regulating the transplacental transport action through the endocytosis pathway. Observe the transplacental transport characterization of PANs in pregnant rat model qualitatively. The study provide the basis for further research and application of PANs in the perinatal medicine field, and provide new ideas to control the transplacental transport behaviour for nano drug delivery system.

药物透过胎盘屏障可能对胎儿产生毒性作用是妊娠期用药面临的重大挑战，纳米药物输送系统透过胎盘屏障的机制及可控策略是纳米药物应用于围产医学亟待研究的问题。申请者前期成功制备了安全性好、对包载药物具有缓释功能的乙酰普鲁兰纳米药物载体（PANs），在此基础上，本项目以人胎盘绒膜癌（BeWo）细胞建立体外胎盘屏障模型，考察异硫氰酸荧光素 (FITC) 标记的纳米粒子PANs-FITC透过胎盘屏障的机制，应用内吞抑制剂对BeWo胎盘屏障模型进行预处理，应用荧光显微镜、透射电镜和激光共聚焦等方法观测PANs的转运，ELISA法检测相关细胞因子水平，阐释PANs透过率-内吞抑制剂-细胞因子水平之间的相关性，明确内吞途径调控PANs透过胎盘屏障的可行性，并用妊娠大鼠模型考察PANs透过胎盘屏障的能力。本课题为PANs在围产医学领域的进一步研究和应用提供实验依据，为纳米递药系统透过胎盘屏障的调控提供新思路。

妊娠期用药十分普遍，药物透过胎盘屏障的特征是妊娠期疾病治疗选择的重要依据，其对胎儿潜在的毒性是妊娠期用药面临的重大挑战。纳米药物输送系统透过胎盘屏障的机制及可控策略是纳米药物应用于围产医学亟待研究的问题。课题组成功制备了安全性好、对包载药物具有缓释功能的乙酰普鲁兰纳米粒子（PANs），在此基础上，将异硫氰酸荧光素 （FITC）对PANs进行荧光标记，成功构建人胎盘绒膜癌（BeWo）细胞模型，考察了纳米粒子PANs-FITC对BeWo细胞的细胞毒性以及透过胎盘屏障的情况，并应用四种内吞抑制剂（氯丙嗪、阿米洛利、制霉素和秋水仙碱）和炎症因子抑制剂（四氢化吡咯二硫代氨基甲酸酯和地塞米松）对BeWo胎盘屏障模型进行预处理，应用流式细胞仪、荧光显微镜、透射电镜和激光共聚焦等方法观测PANs的转运，ELISA法检测相关细胞因子水平。TEER、ZO-1和生物透射电镜结果均可表BeWo细胞呈单层，PA-FITC对BeWo细胞未见明显的细胞毒性，抑制剂均有显著的抑制吞噬和跨BeWo细胞模型转运的作用，细胞因子未见显著的改变，说明了内吞途径和炎症因子抑制剂调控PANs透过胎盘屏障细胞模型的可行性，但是其与炎症因子的相关性有待进一步深入考察。本课题为PANs在围产医学领域的进一步研究和应用提供了实验依据，为研究纳米药物跨胎盘屏障的调控方法提供了新思路。