Zhang Jian, Zhu Xi, Jin Yun, Shan Wei, Huang Yuan
Key Laboratory of Drug Targeting and Drug Delivery System, Ministry of Education, West China School of Pharmacy, Sichuan University , No. 17, Block 3, Southern Renmin Road, Chengdu 610041, P. R. China.
Mol Pharm. 2014 May 5;11(5):1520-32. doi: 10.1021/mp400685v. Epub 2014 Apr 11.
Modifying nanoparticles with targeting peptides which can specifically bind to intestinal epithelium was recently suggested as a strategy to further enhance their ability for the oral delivery of macromolecular drugs. However, few studies were focused on comprehensive understanding of the uptake and transport processes as well as the underlying molecular signaling pathways mediated by the ligand modification. In the present study, the mechanisms of cellular uptake and the tight junction opening associated with the trimethyl chitosan based nanoparticles (M-NPs) and their goblet cell-targeting CSK (CSKSSDYQC) peptide modified nanoparticles (CSK-M-NPs) were investigated. Compared with single ion cross-linked nanoparticles (S NPs), M-NPs and CSK-M-NPs, prepared with multiple agents, exhibited superior stability which could effectively protect drugs against the degradation of trypsin. Caveolae-mediated endocytosis and macropinocytosis were involved in the intracellular uptake of both M-NPs and CSK-M-NPs on Caco-2/HT29-MTX cocultured cells. However, CSK peptide modification could further induce clathrin-mediated endocytosis of the NPs. Intriguingly, most endocytosis subpathways have been altered after CSK peptide modification. Moreover, the opening of epithelial tight junctions was investigated at both protein and gene levels. The results indicated that both M-NPs and CSK-M-NPs could transiently and reversibly open the epithelial tight junctions via the C-Jun NH2-terminal kinase-dependent pathway. However, CSK peptide modification enabled a more rapid opening and recovering of the tight junctions. In all, the enhanced uptake and transport capacity of nanoparticles after CSK peptide modification may be attributed to the alteration of internalization pathways and the stronger ability of opening tight junctions.
最近有人提出用能特异性结合肠上皮的靶向肽修饰纳米颗粒，作为进一步提高其口服递送大分子药物能力的策略。然而，很少有研究专注于全面了解其摄取和运输过程以及由配体修饰介导的潜在分子信号通路。在本研究中，研究了基于三甲基壳聚糖的纳米颗粒（M-NPs）及其杯状细胞靶向CSK（CSKSSDYQC）肽修饰的纳米颗粒（CSK-M-NPs）的细胞摄取机制和紧密连接开放情况。与单离子交联纳米颗粒（S NPs）相比，用多种试剂制备的M-NPs和CSK-M-NPs表现出优异的稳定性，能有效保护药物免受胰蛋白酶降解。小窝介导的内吞作用和巨胞饮作用参与了M-NPs和CSK-M-NPs在Caco-2/HT29-MTX共培养细胞中的细胞内摄取。然而，CSK肽修饰可进一步诱导纳米颗粒的网格蛋白介导的内吞作用。有趣的是，CSK肽修饰后大多数内吞亚途径都发生了改变。此外，还在蛋白质和基因水平上研究了上皮紧密连接的开放情况。结果表明，M-NPs和CSK-M-NPs均可通过C-Jun NH2末端激酶依赖性途径短暂且可逆地开放上皮紧密连接。然而，CSK肽修饰使紧密连接的开放和恢复更快。总之，CSK肽修饰后纳米颗粒摄取和运输能力的增强可能归因于内化途径的改变和更强的开放紧密连接的能力。
