Project Summary Acute kidney injury (AKI) is recognized as a major risk for progressive chronic kidney disease (CKD). However, the mechanism by which AKI leads to fibrogenesis and ultimately to end-stage renal disease (ESRD)2 is not well defined, Due to limited knowledge of the primary signals that drive fibrogenesis, effective therapy for CKD is a major unmet medical need. Our data indicate the novel paradigm that renal denervation can prevent renal fibrosis and inflammation in three renal fibrogenesis models: 5/6 nephrectomy (5/6Nx), unilateral ureteral obstruction (UUO) and ischemic renal injury (IRI). Our data indicate that the renal nerve-derived factor, norepinephrine (NE), signaling via α2-adrenergic receptor (α2-AR) plays a key role in initiating fibrogenesis and inflammation and its inhibition pre-or post-injury can reduce fibrosis by about 70% in these CKD models. This finding is striking, as most experimental strategies targeting a single molecule or a pathway rarely achieve a reduction of fibrosis of more than 50%. Our overall goal of this study is to delineate the mechanisms by which NE signaling via α2-AR induces fibrosis and determine the therapeutic potential of inhibition of the α2-AR subtype or its downstream signaling pathways in preventing renal fibrogenesis and inflammation in the IRI model. Our preliminary studies indicate that NE signaling via α2-AR induces the expression of angiotensinogen (AGT) in renal proximal tubular cell (RPTC) via activation of cAMP-response element-binding protein (CREB). Further, simultaneous inhibition of α2AR subtypes A and C additively protected from inflammation and fibrosis, suggesting activation of subtype specific signaling pathways, parallel to CREB-AGT axis, that may promote interstitial fibrosis and CKD. Based on these data, our central hypothesis is that NE activates α2AR-subtypes specific-signaling pathways to induce interstitial fibrosis and their inhibition can prevent long term sequelae of IRI. Further, α2AR activation regulates parallel pathways including, fibroblast activation and Mφ infiltration, activation and Mφ phenotypic switching, and activates Renin-Angiotensin II Signaling (RAS) signaling pathways to promote fibrogenesis. In specific aim 1, using genetic and pharmacological approaches, we will delineate the functional and mechanistic role of the three α2-AR subtypes (A, B and C), and the effect of their inhibition on renal fibrogenesis in the IRI-model. Using transcriptomic profile, we will identify overlapping versus specific pathways between the α2AR subtypes and identify the signaling molecules that provides added protection after combinatorial inhibition. In specific aim 2, we will dissect out the distinct role of of α2-AR subtype(s) cell-specific role (PTC vs. Mφ vs fibroblasts) in cytokine secretion in fibroblast differentiation, Mφ behavior or Mφ switching and tubular injury. In specific aim 3, we will identify the α2-AR subtype/s that activates CREB and AGT signalin...