Purpose

Many non-viral vectors with positive charge and physical encapsulation of siRNA have been reported. However, the studies of non-viral vectors with chemical conjugation of siRNA and neutral charge were rarely reported.

Methods

A redox-responsive and neutral nanoparticle (NP) was designed using poly(d,l-lactide-co-glycolide) (PLGA)-conjugated siRNA, PLGA, and Chitosan oligosaccharide. The physicochemical properties, stability, permeation ability, redox responsiveness, and in vitro free siRNA release of the NPs were evaluated. In vitro cellular uptake and gene silencing activity of the NPs were also investigated in four cervical cancer cell lines (SiHa, CaSki, HeLa, and c33a).

Results

Redox-responsive and electrically neutral PLGA NPs encapsulating siRNA targeting the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene were formulated. The NPs were modified with chitosan oligosaccharide in order to achieve a neutral surface charge, at a nitrogen/phosphate (N/P) ratio of 20:1, and they had the desired size of ~ 90 nm. They were characterized by a high encapsulation efficiency (approximately 70%), in vitro stability in physiologic conditions, long-term shelf-life stability (> 18 months), and rapid permeation ability through mucus. NP disassembly in reducing environments was rapid. A sustained release phenomenon was observed, which contributed to higher concentrations and prolonged activity of the NPs. The GAPDH silencing efficiency of the NPs was both N/P ratio- and cell type-dependent, and it was as efficient as lipofectamine 2000 in SiHa, CaSki, HeLa, and c33a cells at an N/P ratio of 20:1.

Conclusion

Our findings indicate that redox-responsive and electrically neutral NPs are promising nanocarriers for siRNA delivery to potentially treat human cervical carcinoma.

中文翻译：

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用于人宫颈癌细胞中 siRNA 递送的氧化还原反应和电中性 PLGA 纳米颗粒

目的

已经报道了许多具有正电荷和物理封装 siRNA 的非病毒载体。然而，关于非病毒载体与 siRNA 和中性电荷的化学缀合的研究很少报道。

方法

使用聚 ( d,l-丙交酯-共-乙交酯) (PLGA) 缀合的 siRNA、PLGA 和壳聚糖寡糖设计了一种氧化还原响应性和中性纳米颗粒 (NP)。评估了 NPs 的理化性质、稳定性、渗透能力、氧化还原反应性和体外游离 siRNA 释放。还在四种宫颈癌细胞系（SiHa、CaSki、HeLa 和 c33a）中研究了 NPs 的体外细胞摄取和基因沉默活性。

结果

配制了氧化还原响应性和电中性 PLGA NPs，其封装了靶向 3-磷酸甘油醛脱氢酶 (GAPDH) 基因的 siRNA。NPs 用壳寡糖修饰以获得中性表面电荷，氮/磷酸盐 (N/P) 比为 20:1，并且它们具有约 90 nm 的所需尺寸。它们的特点是高封装效率（约 70%）、生理条件下的体外稳定性、长期保质期稳定性（> 18 个月）和通过粘液的快速渗透能力。还原环境中的 NP 拆卸速度很快。观察到持续释放现象，这有助于提高 NPs 的浓度和延长活性。NPs 的 GAPDH 沉默效率与 N/P 比和细胞类型有关，

结论

我们的研究结果表明，氧化还原反应性和电中性 NPs 是用于 siRNA 递送的有前途的纳米载体，可能治疗人类宫颈癌。