Abstract Acute kidney injury (AKI) is a major health problem, affecting >1.5 million patients in the US each year. AKI is a precursor to chronic kidney disease and often progresses to kidney failure. Currently there are no effective treatments for AKI and therapies are urgently needed. The kidney has an inherent ability to regenerate following injury, raising the possibility that pro-regenerative therapies for AKI can be developed. Renal regeneration is thought to occur by surviving renal tubular epithelial cells (RTECs) undergoing dedifferentiation to a progenitor-like state followed by proliferation and re-differentiation. In addition, M1/'kill' (classic/pro- inflammatory) and M2/'heal' (alternative/pro-repair) type macrophages play opposing roles in promoting tubular regeneration, respectively, and the balance of these subtypes is critical for healthy repair. For poorly understood reasons, the regenerative process can stall, with RTECs arresting in the G2/M phase of the cell cycle and producing pro-fibrotic cytokines. Therefore, finding drugs that promote tubular repair and reduce fibrosis will have a tremendous clinical impact. Towards this goal, we have developed zebrafish and induced pluripotent stem cell (iPSC)-derived human kidney organoids as models to study AKI and we have identified novel pro-regenerative compounds. We discovered a new class of histone deacetylase (HDAC) inhibitors (HDIs), the phenylthiobutanoates (PTBAs), which specifically inhibit HDAC8, a known modulator of retinoic acid (RA) signaling. We have demonstrated that PTBAs promote renal regeneration by increasing proliferation and decreasing G2/M arrest of zebrafish and mouse RTECs, and reduce post-AKI fibrosis in mice. We discovered that these pro-regenerative activities are dependent on RA signaling during AKI. Studies suggest that RA acts on RTECs as well as macrophages to alter M1/M2 switching and reduce RTEC G2/M arrest. Based on these data, we hypothesize that PTBAs promote renal regeneration by inhibiting HDAC8, thereby enhancing RA signaling, which in turn, induces RTEC proliferation and alters M1/M2 macrophage switching. To test these hypotheses, we propose the following Aims: Aim 1. Elucidate the role of Retinoic Acid and HDAC8 in driving PTBA efficacy. The focus is to confirm HDAC8 as the in vivo target of PTBA, explore the effect of PTBA on the RA pathway, and perform an unbiased RNA-Seq screen to identify RTEC genes affected by PTBA treatment. Aim 2. Examine the relationship between PTBA and the immune response in zebrafish. Our data suggests that RA signaling drives M2/'heal' polarization, we favor a model in which PTBA enhances RA signaling in macrophages, thereby promoting M2/'heal'-mediated renal repair. Aim 3. Establish human kidney organoids as a model of AKI and pre-clinical drug testing. We have developed a simple bioreactor- based method for generating, in bulk, human kidney organoids from iPSCs for modeling AKI.