Conclusion
Our results indicate for the very first time that selective tropism, migration, and infiltration of LPMs into extra-peritoneally located lungs was dependent on clodronate-mediated AM depletion. These results further open the possibility of therapeutically utilizing LPMs as delivery vehicles to carry nanoparticle-encapsulated oligonucleotide modalities to potentially address inflammatory diseases, infectious diseases and even cancer.
Methods
C12-200 cationic lipidoid-based nanoparticles were employed to selectively deliver a small interfering RNA (siRNA)-targeting CD-45 labeled with a cyanine 5.5 (Cy5.5) dye to LPMs in vivo via intraperitoneal injection. We utilized a non-invasive optical technique called Diffuse In Vivo Flow Cytometry (DiFC) to then systemically track these LPMs in real time and paired it with more conventional techniques like flow cytometry and immunocytochemistry to initially confirm uptake of C12-200 encapsulated siRNA-Cy5.5 (siRNA-Cy5.5 (C12-200)) into LPMs, and further track them from the peritoneal cavity to the lungs in a mouse model of AM depletion incited by intranasally administered clodronate liposomes. Also, we stained for LPM-specific marker zinc-finger transcription factor GATA6 in harvested cells from biofluids like broncho-alveolar lavage as well as whole blood to probe for Cy5.5-labeled LPMs in the lungs as well as in systemic circulation.
Results
siRNA-Cy5.5 (C12-200) was robustly taken up by LPMs. Upon depletion of lung-resident AMs, these siRNA-Cy5.5 (C12-200) labeled LPMs rapidly migrated to the lungs via systemic circulation within 12-24 h. DiFC results showed that these LPMs intravasated from the peritoneal cavity and utilized a systemic route of migration. Moreover, immunocytochemical staining of zinc-finger transcription factor GATA6 further confirmed results from DiFC and flow cytometry, confirming the presence of siRNA-Cy5.5 (C12-200)-labeled LPMs in the peritoneum, whole blood and BALF only upon clodronate-administration.