Université Lyon 1, UCBL-INRA-EPHE UMR-754, Retrovirus & Comparative Pathology, Lyon, France.
PLoS One. 2012;7(12):e52326. doi: 10.1371/journal.pone.0052326. Epub 2012 Dec 20.
Cell microparticles (MPs) released in the extracellular milieu can embark plasma membrane and intracellular components which are specific of their cellular origin, and transfer them to target cells. The MP-mediated, cell-to-cell transfer of three human membrane glycoproteins of different degrees of complexity was investigated in the present study, using a CHO cell model system. We first tested the delivery of CAR and CD46, two monospanins which act as adenovirus receptors, to target CHO cells. CHO cells lack CAR and CD46, high affinity receptors for human adenovirus serotype 5 (HAdV5), and serotype 35 (HAdV35), respectively. We found that MPs derived from CHO cells (MP-donor cells) constitutively expressing CAR (MP-CAR) or CD46 (MP-CD46) were able to transfer CAR and CD46 to target CHO cells, and conferred selective permissiveness to HAdV5 and HAdV35. In addition, target CHO cells incubated with MP-CD46 acquired the CD46-associated function in complement regulation. We also explored the MP-mediated delivery of a dodecaspanin membrane glycoprotein, the CFTR to target CHO cells. CFTR functions as a chloride channel in human cells and is implicated in the genetic disease cystic fibrosis. Target CHO cells incubated with MPs produced by CHO cells constitutively expressing GFP-tagged CFTR (MP-GFP-CFTR) were found to gain a new cellular function, the chloride channel activity associated to CFTR. Time-course analysis of the appearance of GFP-CFTR in target cells suggested that MPs could achieve the delivery of CFTR to target cells via two mechanisms: the transfer of mature, membrane-inserted CFTR glycoprotein, and the transfer of CFTR-encoding mRNA. These results confirmed that cell-derived MPs represent a new class of promising therapeutic vehicles for the delivery of bioactive macromolecules, proteins or mRNAs, the latter exerting the desired therapeutic effect in target cells via de novo synthesis of their encoded proteins.
细胞微粒（MPs）释放到细胞外环境中，可以携带其细胞起源特有的质膜和细胞内成分，并将其转移到靶细胞。本研究使用 CHO 细胞模型系统，研究了三种不同复杂程度的人类膜糖蛋白通过 MPs 介导的细胞间转移。我们首先测试了 CAR 和 CD46 的传递，这两种单跨膜蛋白分别作为人腺病毒血清型 5（HAdV5）和 35（HAdV35）的高亲和力受体。我们发现，CHO 细胞（MP-供体细胞）组成性表达 CAR（MP-CAR）或 CD46（MP-CD46）的 MPs 能够将 CAR 和 CD46 转移到靶 CHO 细胞，并赋予其对 HAdV5 和 HAdV35 的选择性许可。此外，与 MP-CD46 孵育的靶 CHO 细胞获得了补体调节相关的 CD46 功能。我们还探索了将十二跨膜糖蛋白 CFTR 通过 MPs 递送至靶 CHO 细胞。CFTR 在人细胞中作为氯离子通道，与遗传性疾病囊性纤维化有关。与 CHO 细胞组成性表达 GFP 标记的 CFTR（MP-GFP-CFTR）的 MPs 孵育的靶 CHO 细胞被发现获得了新的细胞功能，即与 CFTR 相关的氯离子通道活性。靶细胞中 GFP-CFTR 出现的时间进程分析表明，MPs 可以通过两种机制实现 CFTR 向靶细胞的递：成熟的、插入质膜的 CFTR 糖蛋白的转移，以及 CFTR 编码 mRNA 的转移。这些结果证实，细胞来源的 MPs 代表了一类有前途的治疗性载体，用于传递生物活性大分子、蛋白质或 mRNA，后者通过其编码蛋白的从头合成在靶细胞中发挥所需的治疗效果。
