Therapeutic cargo properties of isolated exosomes
The cargo properties of the isolated exosomes from different methods were investigated using quercetin-loaded exosome mixtures on pre-cultured and doxorubicin-induced senescent neonatal human dermal fibroblasts. The results show no significant differences in relative β-galactosidase activity between 160μM of free quercetin compared to 80μM quercetin loaded exosomes isolated by SEC (P>0.05) and TEI (P>0.05) (Figure 5), but significantly higher than the quercetin loaded with IP and exosomes alone. This indicates the potential increase (almost doubling) of the intracellular concentration of quercetin loaded in exosomes isolated by SEC and TEI in par with that of 160μM free quercetin. Compared to negative control, all isolated EVs loaded with quercetin significantly increased cell viability (Figure 6). However, quercetin loaded into TEI isolated exosomes showed a significantly higher cell viability percentage than the other two isolation methods. The viability is similar to that of the 160μM free quercetin and that of 150μM resveratrol, a potent antioxidant used as a positive control in this experiment. This finding confirms the concept of exosome’s ability in increasing bioavailability as well as the reliability of TEI as the most suitable exosome extraction method. Therefore, TEI extraction method was selected as the most suitable method and, thus, used for the subsequent in vivo testing.