叶酸修饰的聚乙烯亚胺聚合物载PD-L1 siRNA体外靶向胃癌细胞的研究

发布时间：2019-03-25 10:19

【摘要】：目的:构建高效的siRNA纳米载体靶向SGC-7901胃癌细胞,并下调胃癌表达的程序性死亡配体1(PD-L1)。方法:检测叶酸(FA)-PEG-SS-PEI-SPION纳米载体与siRNA复合后的粒径、电位等表征;体外实验检验siRNA的结合能力、复合物细胞毒性、细胞摄入能力及转染效率;磁共振(MR)成像检测示踪能力;检验胃癌细胞PD-L1下调效应及共培养T细胞的细胞因子水平。结果:N/P比值为10时,FA-PEG-SS-PEI-SPION完全复合siRNA,形成电位为(9.14±0.80)m V、粒径为(116.7±2.5)nm的多聚复合物。靶向组的转染率为(95.06±0.44)%,与非靶向组的(93.87±1.05)%相当;平均荧光强度为1 892.67±81.51,高于非靶向组的1 324.33±186.58(P0.05)。普鲁士蓝染色和激光共聚焦显微镜成像证实了复合物的细胞摄入。体外MR成像验证了聚合物的MR造影成像能力。靶向组PD-L1的mRNA最低相对表达量为9.07%±0.79%,Western blot显示PD-L1的表达显著降低。共培养实验显示IFN-γ和TNF-α的分泌水平增加,IL-10的分泌水平降低(P0.05)。结论:本研究构建了FA-PEG-SS-PEI-SPION纳米载体,并证明了其体外靶向细胞及载siRNA下调PD-L1表达的能力和MR示踪的能力,是一种高效和安全的靶向治疗纳米载体。
[Abstract]:Aim: to construct an efficient siRNA nano-carrier targeting SGC-7901 gastric cancer cells and down-regulate the expression of programmed death ligand 1 (PD-L1) in gastric cancer. Methods: the size and potential of folic acid (FA)-PEG-SS-PEI-SPION nano-carrier combined with siRNA were measured, and the binding ability, cytotoxicity, uptake ability and transfection efficiency of siRNA were tested in vitro. The down-regulation of PD-L1 in gastric cancer cells and the cytokine level of co-cultured T cells were detected by magnetic resonance (MR) imaging. Results: when the N / P ratio was 10:00, the FA-PEG-SS-PEI-SPION-siRNA, complex formed a polymeric complex with a potential of (9.14 卤0.80) m V,) and a particle size of (116.7 卤2.5) nm. The transfection rate of targeting group was (95.06 卤0.44)%, which was similar to that of non-targeting group (93.87 卤1.05)%, and the average fluorescence intensity was 1 892.67 卤81.51, which was higher than that of non-targeting group (1 324.33 卤186.58, P0.05). Prussian blue staining and laser confocal microscope imaging confirmed the cellular uptake of the complex. In vitro MR imaging demonstrated the ability of MR imaging of polymers. The lowest relative expression of mRNA in PD-L1 of target group was 9.07% 卤0.79%. Western blot showed that the expression of PD-L1 was significantly decreased. Co-culture experiment showed that the secretion of IFN- 纬 and TNF- 伪 increased and the secretion of IL-10 decreased (P0.05). Conclusion: in this study, the FA-PEG-SS-PEI-SPION nano-carrier was constructed, and its ability to target cells in vitro, down-regulate the expression of PD-L1 with siRNA and the ability of MR tracer were proved to be an efficient and safe nano-carrier for targeted therapy.
【作者单位】：中山大学孙逸仙纪念医院消化内科;中山大学孙逸仙纪念医院急诊科;中山大学肿瘤防治中心胸外科;
【基金】：广州市科技计划项目(No.201704020121)
【分类号】：R735.2

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