Exosomes are small single-membrane secreted organelles of approximately 30-150 nm in diameter that are secreted by most, if not all cells. There is a growing appreciation that exosomes are an important part of inter-cellular communication and that cargo carried by exosomes transmit a variety of signaling properties from their cell of origin to cells that receive exosome cargo. Further, the exosome cargo released from a specific cell-type may reflect the physiologic state of that particular cell population. Most studies on exosomes rely on isolating exosomes from cultured cells or cell clusters in vitro, or bulk exosome isolation from in vivo sampling of biological fluids (such as plasma). However, there exist no simple tools that allow to in vivo isolate and examine exosomes that can be traced back to a specific cell of origin and which can be sampled longitudinally over time within the same organism. Thus, we have little information on how exosomes and their cargo released from a certain cell changes with ageing during the course of the life of an organism or how the exosome cargo may change if a particular cell-type responds to increased functional demand and/or injury. To address these questions, we have generated a mouse model that permits conditional and cell-selective genetic labeling of exosomes through expression of an engineered exosome surface marker. We have used a cDNA cassette in which the coding sequence of a green-fluorescent tag (pHluorin) is inserted into the first extracellular domain of the tetraspanin CD63, which is highly enriched in and expressed on the surface of exosomes. We have inserted this cassette into the Rosa26 locus allowing it to be conditionally expressed in a cell-specific manner after CRE-mediated removal of upstream transcriptional stop sequences (= R26- StopLoxP-CD63-pHluorin mouse). From plasma of these mice, using an antibody that recognizes pHluorin antigen, we can isolate and capture exosomes that can be traced back to their cell of origin. This unique mouse model represents a significant advance over existing means to study exosomes in vivo in a cell-selective manner. We have chosen to study exosomes released from pancreatic ß-cells, as these cells are known to undergo distinct functional and molecular changes with ageing and under different physiologic and pathologic conditions. In this exploratory proposal we aim to use our model system to examine whether and how the exosome cargo from pancreatic ß-cells in mice changes over their lifetime and during circumstances of altered metabolic demand. At the molecular level we will focus our proof-of-principle studies on examining changes in RNA levels in ß-cells and in circulating ß-cell-derived exosomes. The outcomes of these studies will increase our knowledge on whether exosomes accurately reflect the age and functional state of the pancreatic ß-cell and whether exosomes and their cargo could be utilized as diagnostic tools for early detection of cellu...