In vitro and in vivo analysis of microvesicle-mediated metastasis using a bright, red-shifted bioluminescent reporter protein of extracellular vesicles

Cancer cells produce heterogeneous extracellular vesicles (EVs) as mediators of intercellular communication. Our study focused on a novel method to image EV subtypes and their biodistribution in vivo. Regardless of injection routes, we established that reporter EVs isolated from murine mammary carcinoma cells expressing PalmReNL, which utilizes bioluminescence resonance energy transfer (BRET), localized to the lungs. This new EV reporter allowed highly sensitive EV tracking in vitro and in vivo and enabled us to begin studies to understand the commonalities and functional differences of the EV subtypes. We demonstrated the early appearance of metastatic foci in the lungs of mammary tumor-bearing mice following multiple injections of the microvesicle (MV)-enriched fraction derived from mammary carcinoma cells. In addition, the results we present here show that tumor cell-derived MVs act on distant tissues through upregulating LC3 expression within the lung.