不同细胞系对细胞穿透肽的摄取和机制比较

Comparison of mechanisms and cellular uptake of cell-penetrating peptide on different cell lines

细胞穿透肽(cell-penetrating peptide,CPP)作为一种潜在的药物输送高效转运载体一直得到研究者的广泛关注。本文中采用4种肿瘤细胞系(MCF-7、MDA-MB-231、C6和B16F10)分别摄取异硫氰酸荧光素(fluorescein isothiocyanate,FITC)标记的CPP,观察到CPP入胞,并具有时间和浓度的依赖性,同时发现了C6细胞对CPP的胞吐作用,其胞吐动力学符合零级方程;在低温(4℃)和内吞抑制剂存在条件下探讨了CPP入胞的机制。低温条件对CPP的入胞未产生抑制作用;肝素钠作为细胞表面硫酸糖蛋白受体抑制剂对CPP的入胞有较强抑制作用,肝素组对CPP的摄取只达到对照组的3%～10%;而氯丙嗪、氯喹和N-乙酰基-N-异丙基阿米洛利[5-(N-ethyl-N-isopropyl)-amiloride,EIPA]对CPP的入胞影响不大。本研究表明,CPP穿透细胞没有选择性,即缺乏细胞特异性,但CPP的摄取量与细胞种类有关。硫酸蛋白聚糖的吸附介导在CPP穿透细胞中发挥了重要作用。

Cell-penetrating peptide （CPP） can be used in pharmaceutics as a highly efficient drug delivery transporter. In this study, four tumor cell lines （MCF-7, MDA-MB-231, C6, and B16F10） were used to observe the uptake of fluorescein isothiocyanate （FITC） labeled CPP and the effects of time and concentration of CPP on cell penetration was studied. The CPP exocytosis on C6 cell line was observed, and its exocytosis kinetics was described by zero order equation. In addition, low-temperature condition （4 ℃） and endocytosis inhibitors were utilized to investigate the mechanism of CPP uptake by cells. Low-temperature condition did not show significantly inhibition on CPP uptake. Heparin, a membrane glycoprotein receptor inhibitor, showed strong inhibition effect （only 3%？10% of the control） on CPP uptake. Chlorpromazine, chloroquine and 5-（N-ethyl-N-isopropyl）-amiloride （EIPA） showed little effect on CPP uptake. This study indicated that CPP penetration had little selectivity on cell type, but the amount and rate of CPP penetration into cells were related to the type of cell lines. The adsorption of CPP on cell membrane induced by sulfate proteoglycan plays an important role on CPP penetration.