Project 2 (Koch) SUMMARY/ABSTRACT Studies have shown that the heart can secrete factors including those in extracellular vesicles (EVs)/exosomes that can induce significant effects both locally to myocardial tissue and distantly to other organs. This includes adipose tissue where data from our lab has revealed that the heart can secrete substances that can alter adiposity, especially after a high fat diet (HFD). In fact, specific signals are released after HFD when adrenergic signaling in myocytes is altered by manipulation of G protein-coupled receptor kinase 2 (GRK2) levels and activity as increased myocyte GRK2 leads to a lean phenotype after HFD and myocyte-specific GRK2 inhibition leads to an obese phenotype. This has been recapitulated in vitro with conditioned media experiments from myocytes to cultured differentiating adipocytes. GRK2 regulates β-adrenergic receptors (βARs) in the heart where this pathway is crucial for normal and compromised cardiac contractility and function. GRK2 is up-regulated in the heart after cardiac stress, which includes HFD and increased GRK2 leads to compromised βAR signaling and GRK2 is involved in heart failure (HF) progression. The GRK2-adrenergic signaling axis appears to be important in the endocrine activity of the heart as well since we have uncovered data over the last cycle of this grant that manipulation of GRK2 can have profound and significant effects on EV cargo including microRNAs (miRs). We believe these EV/exosomes secreted from stressed hearts/myocytes with increased GRK2 could participate in cardiac dysfunction locally. In addition to HFD, GRK2 can be increased in the hearts of mice by ischemic injury and hypertrophic stress. Previous data and new data in this proposal not only shows that altering GRK2 in the heart can have profound effects distantly on adipose tissue via what is secreted but also can affect local cardiac cells in a paracrine fashion. In fact, increased cellular GRK2 results in increased GRK2 in secreted EV/exosomes including after ischemic injury in vivo and these EVs have detrimental effects when placed on cultured cardiac cells. Thus, manipulation of GRK2 in myocytes can significantly alter the myocardial secretome that appears to have profound effects both locally and distantly including new data showing changes in fat depots after cardiac injury in a sex-specific manner. Specifically, the Central Hypothesis of this proposal is that cardiac stressors, which alters GRK2 levels and signaling can alter the myocardial secretome including the contents of EV/exosomes, which can significantly affect myocardial injury and repair as well as influence regulation of distant organs, especially adipose. Specific Aims are: [1] To identify factor(s) and elucidate novel mechanisms involved in the communicative signals secreted from GRK2-altered myocardium responsible for regulating systemic adiposity after HFD; [2] To determine the mechanistic role of altered myocyte GRK2 levels on EV/ex...