Abstract 362: Live Tracking Mouse Model for Endogenous Exosomes from Heart

Abstract

Exosomes are emerging new category of messengers communicating among cells, tissues and organs. Understanding the kinetics of exosome communication in vivo are a critical foundation for studying exosome functions and developing exosome based drug-delivery models. Current studies of exosome in vivo trafficking largely rely on the administration of labeled exogenous exosomes. These methods may not fully represent endogenous exosome kinetics due to ex vivo exosome manipulations. Here, we established the first inducible endogenous exosome tracking mouse model that tracks endogenous exosome released from cardiomyocytes in vivo . The ultrasensitive and stable Nano-luciferase (NanoLuc) was selected as a reporter due to its 150-fold stronger signal intensity compared to traditional Firefly and Renilla luciferases, and the longest luminescent half-life amongst all known luciferases. We fused NanoLuc reporter with exosome surface marker CD63 for specific labeling of exosomes. The cardiomyocyte-specific αMHC promoter followed by a loxP-STOP-loxP cassette was engineered for a temporal control of labeled exosomes originating from cardiomyocytes. We crossed this cardiomyocyte-specific transgenic mouse with a tamoxifen-inducible Cre mouse (R26CreERT2) to achieve an inducible expression of the reporter. The exosome labeling and distribution were assessed by a luciferase assay and non-invasive bioluminescent live imaging. As expected, CD63NanoLuc expression was tightly controlled and only detected in cardiomyocytes upon induction. The endogenous exosomes released from cardiomyocytes were labeled and detected in vitro in a cell culture supernatant, and in vivo in the animal plasma. A paracrine uptaken of the Nanoluc-labeled exosomes by cardiac fibroblasts in vivo was demonstrated and quantified. A signature distribution profile of the endogenous exosomes released from cardiomyocytes was exhibited. For the first time, this exosome tracking model enables elucidating the endogenous exosome trafficking pattern, and allows future studies of exosome behavior under different conditions. It provides a useful tool for the exploration of biological functions, mechanisms and clinical applications of exosomes in a broad spectrum of researches.