Beneficial and challenges of exosome application in ischemic heart disease

Fig. 1. Exo biogenesis and secretion routes inside the host cells. Internalized Exos are packed inside the intracellular vesicles named early endosomes. Early endosomes mature into later endosomes and MVBs. Along with these changes, several new ILVs are generated via the invagination of the vesicle membrane. Several biomolecules are sequestrated inside the ILVs. MVBs with numerous ILVs are guided toward lysosomal degradation to fuse cell membranes to release their contents (ILVs) into the extracellular matrix where they are called hereafter Exos

Fig. 2. Different molecular pathways are involved in the generation of ILVs. MVBs with numerous intraluminal ILVs are generated using multiple pathways. To be specific, ESCRT-dependent and ESCRT-independent pathways can control the generation of de novo ILVs. In the canonical ESCRT-dependent pathways, four complexes including ESCRT-0, -I, -II, -III along with ATPase VPS4 regulate the dominant process of ILV formation (1). The complementary non-canonical ESCRT-dependent pathways, effectors such as HD-PTP (2) or Alix (3) in close collaboration with ESCRT-III and VPS4 sequestrate certain biosignaling molecules into the MVBs. In line with these activities, lipid rafts belonging to non-canonical ESCRT-dependent systems are also important. More interestingly, some tetraspanins such as CD63 are involved in the generation of nascent ILVs by both ESCRT and ceramide-independent mechanisms. It is thought that the nSMase2-ceramide pathway is another important molecular axis that regulates the formation of ILVs in MVBs in an ESCRT-independent manner. Factors such as caveolin-1 or flotillins proceed with the formation of ILVs by lipid raft-dependent pathways with simultaneous activation of the…

Fig. 3. Several possible Exo applications for the alleviation of ischemic myocardium. Each approach has its advantages and limitations. The systemic administration of Exos can yield low on-target effects while increasing the rapid elimination of circulating Exos via the activity of immune cells. Local administration of free Exos (direct intracoronary artery, intramyocardial, intrapericardial, and catheter-based intracardial injections) or injection via supporting substrates and hydrogels can increase the on-target delivery efficiency into the ischemic area. Created with BioRender online software

Fig. 4. Different strategies are used for cargo loading and increase of on-target delivery of Exos into the ischemic myocardium. The sophisticated manipulations can be done on the parent cells or directly on the purified Exos before injection into the target sites. These approaches increase the delivery efficiency and therapeutic outcomes of administrated Exos in the ischemic myocardium. Created with BioRender online software

Fig. 5. CRISPR/Cas9-mediated genome modification for the increase of therapeutic efficiency of Exos. Up-regulation, down-regulation, deletion, and or addition of certain factors, or improvement of delivery ligands and delivery molecules can be done in the genetic pool using CRISPR/Cas9. Created with BioRender online software

Fig. 6. Cardiomyocyte-specific binding peptide (CMP)-targeted Exos reduced mouse cardiomyocyte programmed cell death in vitro after 7 days (a–e; Scale bar: 100 µm). Cardiomyocytes were incubated with peptide (WLSEAGPVVTVRALRGTGSW) plus vehicle (10 µM DMSO) (b), naïve Exos (c), CMP-targeted Exos (d), or CMP-targeted Exos + synthetic peptide (e) for 24 h for controlling the apoptosis. Data indicated that cells co-treated with CMP-targeted Exos or CMP-targeted Exos plus synthetic peptide had fewer apoptotic changes. *p < 0.05 (compared with peptide plus vehicle control), and **p < 0.05 compared with Exos and CMP-targeted Exo + peptide. One-way ANOVA followed by Tukey’s Multiple Comparison post-hoc test. [114]. Copyright 2019; Scientific Reports

Fig. 7. Administration of miR-126-loaded CPC EVs (miR-126 + ELVs) in rats with I/R injury led to angiogenesis after 28 days (A–D). Immunofluorescence staining for detection of isolectin-B4+ capillaries, SMA+ arterioles, and MHC+ large vessel (A). Measuring the local density of the isolectin-B4+ area and vessel size (B), SMA+ arterioles (C), and MHC+ large vessel (D) in the myocardium. sEV-like vehicle: ELVs; SMA = smooth muscle actin; Small extracellular vesicles; sEVs; SM-MHC = smooth muscle-myosin heavy chain. Scale bar = 100 μm; One-way ANOVA with Tukey post hoc analysis. n.s. = not significant. *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001. [120]. Copyright 2023; ACS Nano