Project description:Maximizing the potential of human kidney organoids for drug testing, regenerative medicine and to model development and disease requires addressing cell immaturity, the lack of a branching collecting system and the off target cell types. Here we establish methods to independently generate the two kidney progenitor cell populations – metanephric mesenchyme and ureteric bud. Combining these two progenitor cell types results in organoids with an improved branched collecting system. We also identified the hormones aldosterone and arginine vasopressin as critical to promote maturation of collecting duct cell types. The resulting organoids contain the full range of epithelial cells in the nephron, including principal and intercalated cells. By scRNA-seq, we demonstrate superior proximal tubule maturation and reduced off-target cell populations using this protocol.
Project description:Kidney organoids are emerging as an increasingly applied model system to hold great promise for transplantation to individuals with end-stage renal disease and as a useful tool for kidney research. Multiple iPSC-derived renal organoids have been established to understand kidney development and disease. However, few disease models originated from adult renal tissue with fully mature cell fates have been set up to recapitulate kidney inflammation or fibrosis. Here we created a novel organoid model that faithfully representing the cellular state of inflammatory response during the progression of renal disease upon TNFα exposure. scRNA-seq showed signatured inflammatory chemokines, cytokines and inflammation-associated signaling pathways were activated in TNFα-treatment organoids together with injury-associated markers such as LCN2 and CLU. Donor age is associated with TNFα-induced inflammatory reaction in organoids. Kidney organoids from young donors showed more transcriptional changes by down-regulation of TGFβ and extracellular matrix genes. In summary, we established an in vitro TNFα-treated kidney organoid model that representing the cellular state of the inflammatory response during renal disease progression and provided a novel tool for studying inflammation-related kidney disease and drug discovery.
Project description:Kidney organoids are emerging as an increasingly applied model system to hold great promise for transplantation to individuals with end-stage renal disease and as a useful tool for kidney research. Multiple iPSC-derived renal organoids have been established to understand kidney development and disease. However, few disease models originated from adult renal tissue with fully mature cell fates have been set up to recapitulate kidney inflammation or fibrosis. Here we created a novel organoid model that faithfully representing the cellular state of inflammatory response during the progression of renal disease upon TNFα exposure. scRNA-seq showed signatured inflammatory chemokines, cytokines and inflammation-associated signaling pathways were activated in TNFα-treatment organoids together with injury-associated markers such as LCN2 and CLU. Donor age is associated with TNFα-induced inflammatory reaction in organoids. Kidney organoids from young donors showed more transcriptional changes by down-regulation of TGFβ and extracellular matrix genes. In summary, we established an in vitro TNFα-treated kidney organoid model that representing the cellular state of the inflammatory response during renal disease progression and provided a novel tool for studying inflammation-related kidney disease and drug discovery.
Project description:Acute kidney injury (AKI) remains a major global healthcare problem and there is a need to develop human-based models to study AKI in vitro. Towards this goal, we have characterized induced pluripotent stem cell-derived human kidney organoids and their response to cisplatin, a chemotherapeutic drug that induces AKI and preferentially damages the proximal tubule. We found that a single treatment with 50 µM cisplatin induces HAVCR1 and CXCL8 expression, DNA damage (γH2AX) and cell death in the organoids in a dose-dependent manner but greatly impairs organoid viability. DNA damage was not specific to the proximal tubule but also affected the distal tubule and interstitial cell populations. This lack of specificity correlated with low expression of the proximal tubule-specific SLC22A2/OCT2 transporter for cisplatin. To improve viability, we developed a repeated low-dose regimen of 4x 5 µM cisplatin over 7 days and found this causing less toxicity while still inducing a robust AKI response that included secretion of known AKI biomarkers and inflammatory cytokines. This work validates the use of human kidney organoids to model aspects of AKI, with the potential to identify new AKI biomarkers and develop better therapies.
Project description:Kidney diseases including acute kidney injury (AKI) and chronic kidney disease (CKD), which can progress to end stage renal disease (ESRD), are a worldwide health burden. Organ trans-plantation or kidney dialysis are the only effective available therapeutic tools. Therefore, in vitro models of kidney diseases and the development of prospective therapeutic means are highly demanded. Within the kidney, the glomeruli are involved in blood filtration and waste excre-tion, and are easily affected by changing cellular conditions. Puromycin aminonucleoside (PAN) is a nephrotoxin, which can be harnessed for imitating acute glomerular damage and modelling of glomerular disease. For this reason, we generated kidney organoids from three iPSC lines and treated these with PAN in order to induce kidney injury. Morphological observations re-vealed disruption of glomerular and tubular structures within the kidney organoids upon PAN treatment, which were confirmed by transcriptome analyses. Subsequent analyses revealed an upregulation of immune response as well as inflammatory and cell death-related processes. We concluded that treatment of iPSC-derived kidney organoids with PAN induces kidney injury mediated an intertwined network of inflammation, cytoskeletal re-arrangement, DNA damage, apoptosis and cell death. Furthermore, urine stem cell-derived kidney organoids can be used for modelling of kidney-associated diseases and drug discovery.
Project description:Current renal organoid models derived from embryonic or induced pluripotent stem cells mimic development. Yet, few studies have attempted to generate organoids from human adult kidney to recapitulate regeneration or pathological dysregulation in vitro. Here, we report a novel expanding regenerative organoids culture system from renal cortex and medulla. Transcriptomic sequencing and immunostaining identified that these organoids share similar molecular features with kidney injury-responsive regeneration. Heterogeneous populations in organoids including cycling epithelial progenitors and differentiated cell types were identified by single cell sequencing including proximal tubules, principal cells and collecting duct (CD) progenitors that can be induced into functional CD system. Furthermore, we established polycystic organoids derived from patients that represent an advanced platform for polycystic kidney disease (PKD) modeling. By drug screening, QNZ, GSK2193874 and AMPK activators were shown to significantly reduce polycystic growth. Our results demonstrated a novel in vitro renal organoid model to study regenerating adult renal cells and PKD mechanism, providing tools for discovery of therapeutic targets.