PROJECT SUMMARY/ABSTRACT Renal cysts are the hallmark of autosomal dominant polycystic kidney disease (ADPKD; the fourth major cause of end-stage kidney disease). The current paradigm of the disease pathogenesis is based on progressive growth of renal cysts over the life of an ADPKD patient, with the most prominent cysts being those that emerged in utero. Data obtained with our new image analysis tools, new animal model, and ADPKD patients show instead that individual cysts' growth rates are highly variable and change over time. In fact, the proportion of growing cysts is nearly matched by those that are regressing, and an index that quantifies this growing and regressing cyst phenotype predicts better renal outcomes than total kidney volume (TKV; an FDA-approved ADPKD biomarker). The use of this new individual cyst-based biomarker in future studies is likely to transform ADPKD classification and staging to achieve earlier and more accurate identification of high-risk patients with ADPKD, including children (who have relatively small TKVs). However, fundamental questions for understanding disease pathobiology and developing new ADPKD therapeutics remain unanswered. For example, why do some cysts grow faster than others, and why do some disappear? Since these questions cannot be answered in mouse models (they require two affected PKD gene alleles to develop cysts and their cysts appear to grow progressively over time without regression), we propose to study these concepts in our new Pkd2K874A*1/+ rat that develops the growing and regressing cysts as a heterozygote (like humans). However, since the cyst growth rates are variable and change over longer time intervals, we propose to use acute kidney injury (AKI) as a model for the initial dynamic phase of individual cyst growth and the later phase of cyst regression. Since complement C3 is one of few factors that modulate AKI and cystogenesis and were associated with the rate of human ADPKD progression, our central hypothesis is that C3 regulates the injury-induced dynamic growth phase of individual cyst growth in carriers of pathogenic ADPKD gene variants. We will address this central hypothesis in two Aims: Aim 1. Test the hypothesis that the injury-induced dynamic growth phase of most renal cysts in heterozygous carriers of the Pkd2 gene defect is mediated by C3 activity - by testing the effects of ischemia-reperfusion injury (IRI) in animals generated from crosses of the available Pkd2K874A*1/+ and C3 knockout rats. We will complement our advanced MRI imaging techniques (allowing individual renal cyst monitoring through their co-registration over time) with a single cell and spatial transcriptomics to reveal which cells and pathways define the dynamic vs. regression phase of individual cyst growth, and how they are affected by C3 deficiency. Aim 2 will replicate Aim 1 in the already available Pkd1 knockout rat model (PKD1 and PKD2 genes are the two major ADPKD genes and their protein products for...