Mononuclear phagocyte system blockade improves therapeutic exosome delivery to the myocardium
Mononuclear phagocyte system blockade improves therapeutic exosome delivery to the myocardium
复制标题
单核吞噬细胞系统阻断可改善治疗性外泌体向心肌的递送
DOI：
10.7150/thno.38198
复制
发表时间：
2020-01-01
期刊：
影响因子：
12.4
通讯作者：
Liu, Li
中科院分区：
文献类型：
--
作者：
Wan, Zhuo;Zhao, Lianbi;Liu, Li
理由：外泌体正在成为一种有前途的药物输送载体。然而，单核吞噬细胞系统（MPS）对外泌体的快速摄取仍然是药物输送到其他靶器官（包括心脏）的障碍。我们假设，通过 MPS 预先阻断外泌体的摄取将改善其向靶器官的递送。方法：从细胞培养基中分离外泌体。通过荧光成像在体外和体内追踪荧光标记的外泌体。通过 qPCR 分析了各种细胞系和器官中网格蛋白重链 (Cltc)、cavolin1、Pak1 和 Rhoa（已知的内吞作用基因）的表达情况。通过Western blotting分析siRNA对Cltc的敲低效率。外泌体（对照）和外泌体（阻断）是通过电穿孔用siControl或siClathrin封装分离的外泌体构建的，而外泌体（治疗）是通过用miR-21a封装分离的外泌体构建的。采用阿霉素诱导的心脏毒性模型，通过超声心动图验证基于外泌体的 miR-21a 递送的治疗效率。结果：外泌体优先在肝脏和脾脏中积累，主要是由于存在丰富的巨噬细胞。除了众所周知的吞噬作用外，有效的内吞作用也有助于巨噬细胞对外泌体的摄取。发现与其他内吞作用相关基因相比，Cltc 在巨噬细胞中高度表达。因此，敲低 Cltc 显着降低体外和体内巨噬细胞对外泌体的摄取。此外，预先注射外泌体（阻断）显着提高了外泌体向脾脏和肝脏以外的器官的递送效率。一致地，与直接注射外泌体（治疗性）相比，预先注射外泌体（阻断性）在阿霉素诱导的心脏毒性小鼠模型中对心脏功能产生了更好的治疗效果。结论：用外泌体（阻断性）预先阻断巨噬细胞对外泌体的内吞作用，成功有效地改善了后续外泌体（治疗性）在靶器官（如心脏）中的分布。已建立的两步外泌体递送策略（首先阻止外泌体的摄取，然后递送治疗性外泌体）将是一种有前途的基因治疗方法。
Rationale: Exosomes are emerging as a promising drug delivery carrier. However, rapid uptake of exosomes by the mononuclear phagocyte system (MPS) remains an obstacle for drug delivery into other targeted organs, including the heart. We hypothesized that prior blocking of uptake of exosomes by the MPS would improve their delivery to the targeted organs.Methods: Exosomes were isolated from the cell culture medium. Fluorescence-labeled exosomes were tracked in vitro and in vivo by fluorescence imaging. The expression of clathrin heavy chain (Cltc), cavolin1, Pak1 and Rhoa, known genes for endocytosis, were profiled in various cell lines and organs by qPCR. The knockdown efficiency of siRNA against Cltc was analyzed by Western blotting. Exosome(control) and exosome(blocking) were constructed by encapsulating isolated exosomes with siControl or siClathrin via electroporation, while exosome(therapeutic) was constructed by encapsulating isolated exosomes with miR-21a. Doxorubicin-induced cardiotoxicity model was used to verify the therapeutic efficiency of the exosome-based miR-21a delivery by echocardiography.Results: Exosomes were preferentially accumulated in the liver and spleen, mainly due to the presence of abundant macrophages. Besides the well-known phagocytic effect, efficient endocytosis also contributes to the uptake of exosomes by macrophages. Cltc was found to be highly expressed in the macrophages compared with other endocytosis-associated genes. Accordingly, knockdown of Cltc significantly decreased the uptake of exosomes by macrophages in vitro and in vivo. Moreover, prior injection of exosome(blocking) strikingly improved the delivery efficiency of exosomes to organs other than spleen and liver. Consistently, compared with the direct injection of exosome(therapeutic), prior injection of exosome(blocking) produced a much better therapeutic effect on cardiac function in the doxorubicin-induced cardiotoxicity mouse model.Conclusions: Prior blocking of endocytosis of exosomes by macrophages with exosome(blocking) successfully and efficiently improves the distribution of following exosome(therapeutic) in targeted organs, like the heart. The established two-step exosome delivery strategy (blocking the uptake of exosomes first followed by delivery of therapeutic exosomes) would be a promising method for gene therapy.