Protein-loaded soluble and nanoparticulate formulations of ionic polyphosphazenes and their interactions on molecular and cellular levels
Protein-loaded soluble and nanoparticulate formulations of ionic polyphosphazenes and their interactions on molecular and cellular levels
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DOI：
10.1016/j.msec.2019.110179
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发表时间：
2020-01-01
影响因子：
7.9
通讯作者：
Fuerst, Thomas R.
中科院分区：
文献类型：
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作者：
Andrianov, Alexander K.;Marin, Alexander;Fuerst, Thomas R.
通过离子型聚磷腈和蛋白质货物-溶菌酶（LYZ）在接近生理pH（pH 7.4）的水溶液中在存在和不存在离子型交联剂-精胺四盐酸盐的情况下的自组装来制备离子型聚磷腈和蛋白质货物-溶菌酶（LYZ）的纳米颗粒和水溶性制剂。使用不对称流场流分级（AF 4）和动态光散射（DLS）方法研究了LYZ包封效率，制剂的理化特性和反应参数的影响。使用可溶性寡糖底物评价两种聚合物制剂对包封的LYZ的影响，而通过测量包封的LYZ对溶壁微球菌细胞的酶活性来评估它们将蛋白质呈递至细胞表面的能力。发现可溶性和交联的聚合物基质都减少了LYZ对细菌细胞的裂解，而包封的蛋白质对寡糖底物的活性几乎保持不变，表明聚磷腈对蛋白质完整性没有不利影响。此外，当与它们的可溶性对应物相比时，纳米颗粒制剂在细胞测定中显示出明显不同的行为。与水溶性LYZ-PCPP制剂相比，包封在聚磷腈纳米颗粒中的LYZ在其裂解细胞的能力方面显示出约2.5倍的活性。还开发了聚磷腈纳米颗粒的聚乙二醇化的新方法。该方法利用了一种新的离子聚磷腈衍生物，其中包含接枝（聚乙二醇）链。PEG化允许对纳米颗粒的尺寸的改进的控制和对它们的交联密度的更广泛的调节，同时仍然允许蛋白质呈递到细胞基质。
Nanoparticulate and water-soluble formulations of ionic polyphosphazenes and protein cargo - lysozyme (LYZ) were prepared by their self-assembly in aqueous solutions at near physiological pH (pH 7.4) in the presence and absence of an ionic cross-linker - spermine tetrahydrochloride. Efficiency of LYZ encapsulation, physicochemical characteristics of formulations, and the effect of reaction parameters were investigated using asymmetric flow field flow fractionation (AF4) and dynamic light scattering (DLS) methods. The effect of both polymer formulations on encapsulated LYZ was evaluated using soluble oligosaccharide substrate, whereas their ability to present the protein to cellular surfaces was assessed by measuring enzymatic activity of encapsulated LYZ against Micrococcus lysodeikticus cells. It was found that both soluble and cross-linked polymer matrices reduce lysis of bacterial cells by LYZ, whereas activity of encapsulated protein against oligosaccharide substrate remained practically unchanged indicating no adverse effect of polyphosphazene on protein integrity. Moreover, nanoparticulate formulations display distinctly different behavior in cellular assays when compared to their soluble counterparts. LYZ encapsulated in polyphosphazene nanoparticles shows approximately 2.5-fold higher activity in its ability to lyse cells as compared with water-soluble LYZ-PCPP formulations. A new approach to PEGylation of polyphosphazene nanoparticles was also developed. The method utilizes a new ionic polyphosphazene derivative, which contains graft (polyethylene glycol) chains. PEGylation allows for an improved control over the size of nanoparticles and broader modulation of their cross-linking density, while still permitting for protein presentation to cellular substrates.