Currently, therapeutic angiogenesis via direct delivery of growth factors has shown great promise in animal models, but has not been successful in humans thus far. Therefore, continuing search for effective therapeutic approaches aimed at promoting myocardial angiogenesis is desperately needed. Our recent work interestingly showed that Hsp20-overexpressing cardiomyocytes (Hsp20-myocytes) possessed pro- angiogenic capacity, which was associated with increased secretion of exosomes, a group of naturally occurring nano-vesicles (30-100nm) actively released from cells. In addition, our latest data showed that: 1) exosomes derived from Hsp20-myocytes, but not from Hsp20-knockdown myocytes, contained a large amount of Hsp20, VEGFR2 and p-Akt, which were efficiently transferred into endothelial cells, resulting in activation of the pro-angiogenic signaling pathway; 2) Hsp20 interacted with Tsg101, a protein known to be involved in exosome biogenesis; and 3) knockdown of Tsg101 by siRNA attenuated exosome secretion from Hsp20-myocytes. Thus, it will be very important to test whether exosome quantity (Tsg101-dependent exosome biogenesis) or quality (Hsp20-reprogrammed exosomes) is critical for Hsp20-myocyte-elicited pro- angiogenesis. We hypothesize that exosomes and Hsp20 are both required for cardiomyocyte-catalyzed angiogenesis, and that Hsp20-reprogrammed exosomes have therapeutic benefits to diabetic animal hearts, which exhibit an impaired post-ischemic angiogenesis and microvascular rarefaction. These hypotheses will be tested by pursuing three specific aims: 1) Determine whether exosomes are essential for Hsp20- myocyte-induced angiogenesis. Currently, we are generating a mouse model with cardiac-specific deletion of Tsg101 to inhibit exosome production. This model will be crossed with Hsp20-transgenic mice for testing whether Hsp20-myocyte-elicited pro-angiogenic effects are attenuated by blockade of exosome generation. 2) Determine whether Hsp20 is required for exosome-mediated myocardial angiogenesis. We are creating an inducible mouse model with cardiac-specific deletion of Hsp20 to test whether Hsp20 plays an essential role in reprogramming pro-angiogenic exosomes. 3) Test whether Hsp20-stem cell- derived exosomes improve angiogenesis in diabetic animal hearts. We propose to use bone marrow- derived mesenchymal stem cells as a therapeutic source of Hsp20-reprogrammed exosomes, because they are more readily obtained than cardiomyocytes. We will use streptozotocin (STZ)-induced diabetic rats and the Goto-Kakizaki (GK) rats, two well-established diabetic animal models, to pursue this translational research. Together, the proposed studies are expected to unveil a previously unrecognized role of Hsp20 in exosomal reprogramming and in the promotion of cardiomyocyte-induced angiogenesis. Additionally, it is expected to provide novel insights that lead to the development of original exosome-based therapeutic strategies for the treatment of coronary artery disease.