A Human Organoid Model of Polycystic Kidney Disease

多囊肾病的人类器官模型

A HUMAN ORGANOID MODEL OF POLYCYSTIC KIDNEY DISEASE ! PROJECT SUMMARY Polycystic kidney disease (PKD) is the world's most common life-threatening genetic disease and fourth-leading cause of kidney failure, affecting approximately 12 million people worldwide. In PKD, the normal tubular architecture of the kidneys and other organs is gradually replaced by cysts and fibrosis. There is no cure for PKD, and candidate therapeutics are of uncertain efficacy and safety. PKD is commonly inherited as a germline heterozygous loss-of-function mutation in PKD1 or PKD2, encoding polycystin-1 (PC1) and polycystin-2 (PC2), respectively. These large, transmembrane proteins form a channel-receptor complex at the primary cilium and other sites. It is not yet known how mutations result in cyst formation from tubular epithelial cells. A major barrier to deciphering PKD mechanistically is the lack of experimentally accessible models that faithfully recapitulate PKD- specific cystogenesis from tubules. Primary and immortalized cell lines are heterogenous, de- differentiated, non-human, or represent only later stages of disease, while animal models differ substantially from humans and are challenging to decipher mechanistically. To overcome this gap, our laboratory is pioneering the use of human pluripotent stem cells (hPSC) for modeling PKD. hPSC represent a very early embryonic state and provide a renewable source of patient-matched human cells for analysis and regeneration. We have established techniques to differentiate hPSC into human kidney organoids, which are complex, multicellular structures with patterned segments that resemble nephrons. We have further compared gene-edited and patient-derived organoids to mouse and human tissue samples with disease to complement and validate this new system. In hPSC with PKD mutations, we have identified several disease-relevant phenotypes, including cystogenesis from kidney organoid tubules. The goal of this proposal is to elucidate the mechanistic determinants of human PKD cystogenesis in kidney organoids. Based on our preliminary data, we hypothesize that balanced expression of PC1 and PC2 regulates physical adhesion between the kidney tubule and its microenvironment. In Aim 1, we will establish a more faithful experimental model of human PKD by quantifying cystogenesis in heterozygous kidney organoids. In Aim 2, we will investigate how polycystin protein levels are controlled by clarifying the mechanisms underlying PC1 loss in PKD2-/- cells. Finally, Aim 3 will explore a novel hypothesized molecular pathway for PKD by identifying defects in cell adhesion during cyst initiation. Key findings will be validated in primary PKD tissues and non-organoid cells. Collectively, these studies will unveil critical molecular pathways that underlie the enigmatic process of cyst formation, revealing new potential targets for therapeutic intervention.