size-exclusion chromatography, centrifugation and ultrafiltration [36–38]. As EVs
isolated by ultracentrifugation tended to be aggregated more [39], in vivo distribution of EVs isolated using this method should be different from those obtained
using other methods. Furthermore, different drug/biomaterial loading methods of
EVs would have also changed the physiology of EVs [40]. As aforementioned in
the previous section, loading methods can be roughly divided into two categories:
manipulating parent cells followed by isolating EVs and isolation of EVs followed
by loading materials. Drug-loaded EVs prepared by these two different methods
would have resulted in the differences of the characteristics of EVs even though
they were derived from the same cell sources [40]. Direct comparison between
these two methods was performed [18]. Directly labeled EVs with fluorescence dye
showed higher accumulation in the liver, while EVs collected from
fluorescence-labeled parent cells showed higher accumulation in the injured tissue.
When electroporation was used to load nucleic acids in EVs [28, 41], electroporation itself cause aggregation of EVs [28]. Thus, the results of EVs’ biodistribution
using this labeling method for magnetic nanoparticles should be carefully interpreted and even we need to suspect that labeled EVs cannot represent the physiology of unlabeled EVs [26].
In vivo administration methods also influence the biodistribution as well as the
amount of the administered dose of EVs. Wiklander et al. investigated systemically
the effects of administration routes, cell source and targeting on the biodistribution
pattern [35]. For example, intravenously administered EVs resulted in
dose-dependent decrease in the accumulation in the liver and increase in the
accumulation in the bowel. Saturation of the mononuclear phagocyte system of the
liver was proposed as the mechanism. This dose-dependence in biodistribution
pattern is important when we try to use imaging to trace the EVs’ biodistribution to
represent the biodistribution of therapeutic dose of EVs, as a preliminary imaging
study using tracer amount of EVs may not represent biodistribution of later use of
therapeutic dose of EVs. Keeping this in mind, one can try the simultaneous
imaging and therapeutic trial of using EVs [42]. In this so-called theranostic
approach, we mix (radio-)labeled tracer EVs with therapeutic amount of EVs and
inject them and trace the biodistribution. Of course the labeling should be done in
very mild and physiologic condition minimizing the effect of labeling procedure not
to change the characteristics of EVs.
Another factor affecting the in vivo biodistribution is the administration route.
EVs derived from melanoma cells of different studies showed distinctive accumulation patterns according to the administration route [43]. Subcutaneously
injected melanoma-derived EVs accumulated in the lymph nodes [11, 26], while
intravenously injected melanoma-derived EVs mainly accumulated in bone marrow
and lungs [34]. Or another study showed that the major accumulated organ of the
melanoma-derived EVs was liver and spleen when they were injected intravenously
[10]. Head-to-head comparison of EVs derived from the same cell line (HEK293T)
was investigated later. When the three administration routes, intravenous,
intraperitoneal and subcutaneous administration, were compared, intraperitoneal
and subcutaneous injection resulted in lower accumulation in the liver and spleen
8 Endogenous Radionanomedicine: Biodistribution and Imaging
159
isolated by ultracentrifugation tended to be aggregated more [39], in vivo distribution of EVs isolated using this method should be different from those obtained
using other methods. Furthermore, different drug/biomaterial loading methods of
EVs would have also changed the physiology of EVs [40]. As aforementioned in
the previous section, loading methods can be roughly divided into two categories:
manipulating parent cells followed by isolating EVs and isolation of EVs followed
by loading materials. Drug-loaded EVs prepared by these two different methods
would have resulted in the differences of the characteristics of EVs even though
they were derived from the same cell sources [40]. Direct comparison between
these two methods was performed [18]. Directly labeled EVs with fluorescence dye
showed higher accumulation in the liver, while EVs collected from
fluorescence-labeled parent cells showed higher accumulation in the injured tissue.
When electroporation was used to load nucleic acids in EVs [28, 41], electroporation itself cause aggregation of EVs [28]. Thus, the results of EVs’ biodistribution
using this labeling method for magnetic nanoparticles should be carefully interpreted and even we need to suspect that labeled EVs cannot represent the physiology of unlabeled EVs [26].
In vivo administration methods also influence the biodistribution as well as the
amount of the administered dose of EVs. Wiklander et al. investigated systemically
the effects of administration routes, cell source and targeting on the biodistribution
pattern [35]. For example, intravenously administered EVs resulted in
dose-dependent decrease in the accumulation in the liver and increase in the
accumulation in the bowel. Saturation of the mononuclear phagocyte system of the
liver was proposed as the mechanism. This dose-dependence in biodistribution
pattern is important when we try to use imaging to trace the EVs’ biodistribution to
represent the biodistribution of therapeutic dose of EVs, as a preliminary imaging
study using tracer amount of EVs may not represent biodistribution of later use of
therapeutic dose of EVs. Keeping this in mind, one can try the simultaneous
imaging and therapeutic trial of using EVs [42]. In this so-called theranostic
approach, we mix (radio-)labeled tracer EVs with therapeutic amount of EVs and
inject them and trace the biodistribution. Of course the labeling should be done in
very mild and physiologic condition minimizing the effect of labeling procedure not
to change the characteristics of EVs.
Another factor affecting the in vivo biodistribution is the administration route.
EVs derived from melanoma cells of different studies showed distinctive accumulation patterns according to the administration route [43]. Subcutaneously
injected melanoma-derived EVs accumulated in the lymph nodes [11, 26], while
intravenously injected melanoma-derived EVs mainly accumulated in bone marrow
and lungs [34]. Or another study showed that the major accumulated organ of the
melanoma-derived EVs was liver and spleen when they were injected intravenously
[10]. Head-to-head comparison of EVs derived from the same cell line (HEK293T)
was investigated later. When the three administration routes, intravenous,
intraperitoneal and subcutaneous administration, were compared, intraperitoneal
and subcutaneous injection resulted in lower accumulation in the liver and spleen
8 Endogenous Radionanomedicine: Biodistribution and Imaging
159