Mononuclear phagocyte system blockade improves therapeutic exosome delivery to the myocardium

Abstract

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. Exosomecontrol and exosomeblocking were constructed by encapsulating isolated exosomes with siControl or siClathrin via electroporation, while exosometherapeutic 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 exosomeblocking strikingly improved the delivery efficiency of exosomes to organs other than spleen and liver. Consistently, compared with the direct injection of exosometherapeutic, prior injection of exosomeblocking 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 exosomeblocking successfully and efficiently improves the distribution of following exosometherapeutic 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.

Advances in Exosomes‐Based Drug Delivery Systems

“…This approach has been applied to drugs [ 117 ] though it has mainly been used to RNA incorporation into vesicles. [ 118–120 ] This process also leads to exosomes slightly larger than the initial ones. [ 117 ] Although this method may cause RNA aggregation it has been used in clinical protocols.…”

<p>Advances in Exosome-Based Drug Delivery and Tumor Targeting: From Tissue Distribution to Intracellular Fate</p>

“… 58 Clearance of exosome is primarily mediated by the mononuclear phagocyte system (MPS). 59 Exosomes carry a slightly negative charge on the surface, and are expected to reduce MPS-mediated clearance compared with positively-charged NPs. 60 However, phosphatidylserine, a negatively charged lipid, is involved in macrophage uptake of exosomes as part of exosome clearance.…”

Targeted delivery of extracellular vesicles in heart injury

“…Wan and colleagues were the first to show that CLTC plays a significant role in EVs uptake by the MPS. Inhibition of CLTC with siRNA was shown to significantly block endocytosis mediated by the MPS in the spleen and liver, thereby increasing the delivery of intravenously injected EVs in the heart 117 . In the therapeutic anticancer area, Belhadj et al .…”