PLGA-TPGS 负载α-TIF-siRNA 纳米粒对疱疹病毒1的抑制作用

摘要

目的：研究以 PLGA-TPGS 生物可降解材料为载体包载α-TIF-siRNA 的纳米粒对 HSV1的抑制作用。方法以 PLGA-TPGS 为载体，采用双乳蒸发法制备包载α-TIF-siRNA 的 PLGA-TPGS 纳米粒（命名为 PLGA-TPGS/α-TIF-siRNA NPs），并对其进行表征，包括粒径大小、zeta 电位、包封率和释放率，用 MTT 法检测纳米粒对上皮细胞和HeLa 细胞的细胞毒作用，免疫荧光观察纳米粒在细胞内的释放，用空斑实验研究体外研究纳米粒对 HSV1病毒的抑制作用。结果 PLGA-TPGS/α-TIF-siRNA NPs 的粒径大小为（257±2.94）nm，zeta 电位为（-31.25±1.70）mV，siRNA的包封率为（56.23±3.68）％，纳米释放 siRNA 呈双相，即在96 h 释放达到50％，之后呈缓慢释放，用 MTT 法分析 PL-GA-TPGS/α-TIF-siRNA NPs 对原代角质形成细胞和 HeLa 细胞几乎无细胞毒性。荧光显微镜能观察纳米粒 siRNA 细胞内释放。PLGA-TPGS/α-TIF-siRNA NPs 能明显延长抑制感染 HeLa 细胞的 HSV1。结论 PLGA-TPGS 纳米粒可以作 siRNA 的载体。PLGA-TPGS/α-TIF-siRNA NPs 在体外对 HSV1病毒具有明显的抑制作用，可以成为治疗 HSV1-诱导的角膜炎在内的相关疾病的候选药物。%Objective To investigate the inhibitory effects of alpha-TIF siRNA-loaded PLGA-TPGS nanoparticles on herpesviruses 1(HSV1). Methods The novel biodegradable PLGA-TPGS nanoparticles(NPs)were prepared as a delivery system of small interfering ribonucleic acid(siRNA)targetingα-TIF for inhibiting HSV1,and its characteristics including size,zeta potentials,entrapment rate and in vitro release rate were represented. The cytotoxicity of α-TIF siRNA-loaded PLGA-TPGS nanoparticles on epithelial cells and HeLa cells was examined by MTT assay. The inhibitory effect of thenanoparticleson HSV1 was detected by barren spot assay. Results The size,zeta potential of PLGA-TPGS/ α-TIF-siRNA NPs was(257 ± 2. 94)nm and(31. 25 ± 1. 70)mV,respectively. The entrapment rate of siRNA was(56. 23 ± 3. 68)% ,and the in vitro release of NPs displayed biphase kinetics,that is,which reached 50% on 96 hours,then,it was slowly released. The cytotoxicity of PLGA-TPGS/ α-TIF-siRNA NPs on primary keratinocytes or HeLa cells was not observed by using MTT assay. The release of SiRNA located in NPs was observed by using fluorescent microscope. PLGA-TPGS/ α-TIF-siRNA NPs could inhibit HSV1 replication in HeLa cells. Conclusion PLGA-TPGS nanoparticles can be used as siRNA carrier. PLGA-TPGS/ α-TIF-siRNA NPs can inhibit HSV1 replication in vitro,which may become candidate drugs for HSV1 infection- related diseases including keratitis.