Lab-Grown Kidney Tubules Offer New Window on Polycystic Kidney Disease

Scientists in Bonn have re-engineered lab-grown human kidney tubules so that they can, for the first time in this system, be pushed to form new cysts on demand — a step that turns a long-standing modelling frustration into a workable platform for studying autosomal dominant polycystic kidney disease, or ADPKD. The work, published in BMC Molecular and Cell Biology, describes two distinct three-dimensional culture methods for so-called tubuloids, miniature epithelial structures grown from adult kidney tissue, and shows that only one of them reliably develops fresh, measurable cysts when a key disease-linked signalling pathway is activated.

Tubuloids are derived from adult stem cells found in kidney tubular tissue and have attracted attention because they grow quickly, remain genetically stable, and consist exclusively of kidney epithelial cells. That focus is well suited to polycystic kidney disease, in which cysts arise from the tubule epithelium itself and progressively disrupt surrounding tissue until kidney function fails. The problem, as the researchers led by Jana Rohe, Arsila Palliyulla Kariat Ashraf and Marieta Ioana Toma report, is that conventional tubuloids already look cystic: they are spherical, fluid-filled sacs with a central lumen, so an experimentally induced cyst is hard to distinguish from the structure’s baseline shape.

To get around this, the team grew tubuloids from tumour-adjacent, non-cancerous tissue of seven patients who underwent kidney surgery at University Hospital Bonn, embedding freshly isolated cells into collagen domes, the standard approach. They then took a second step: dissociating those established tubuloids into single cells and re-seeding them in suspension dishes coated to prevent attachment, with no extracellular matrix scaffold at all. The choice of scaffold mattered enormously. Fresh kidney cells placed directly into suspension never formed tubuloids at all, merely aggregating into loose clusters, confirming that the matrix is essential for initiating the organoids. But once tubuloids had already formed in collagen, their descendants could self-organize in suspension into dense, multi-layered structures with small or no lumen — a fundamentally different architecture.

Bulk RNA sequencing of tubuloids from four donors, cultured across passages three to five, revealed 6,804 genes differentially expressed between the two culture types, with 3,478 upregulated and 3,326 downregulated in suspension versus dome tubuloids. Immunohistochemistry confirmed that both cultures expressed the kidney epithelial markers PAX8, cytokeratin 7 and cytokeratin 19, and neither expressed markers of endothelium or interstitium, meaning the cultures faithfully represent only the kidney epithelium. Both expressed a mixture of nephron segment markers — collecting duct markers such as GATA3, AQP3 and CA2, the proximal tubule marker PDZK1IP1, and the loop of Henle marker CLDN3 — with GATA3-positive nuclei appearing in 50 to 70 percent of cells, suggesting collecting duct-like cells dominate but do not fully compose the tubuloids. Occasional CD13-positive cells with hints of a brush border pointed to rare proximal tubule cells.

The most striking molecular contrast lay in maturation state. Dome tubuloids expressed dedifferentiation and regeneration markers including CD24, VIM, CD44 and SOX9, and their enriched gene sets pointed toward cell proliferation and mitosis — a profile the authors interpret as recapitulating kidney injury repair. Suspension tubuloids, by contrast, strongly expressed genes for the Na+/K+-ATPase pump and other ion channels, hallmarks of functional mature kidney epithelium, and their enriched pathways involved ion transport across the plasma membrane. Notably, both culture types lacked CDH16, the kidney-specific cadherin of the distal nephron, and neither expressed podocyte markers. The collecting duct emphasis is relevant to disease biology: the collecting duct has long been considered the predominant site of cyst origin in ADPKD patients.

The pivotal experiment involved forskolin, a drug that activates adenylyl cyclase and thereby raises intracellular cyclic AMP, or cAMP. Elevated cAMP signalling is a central player in ADPKD: the polycystin-1/polycystin-2 protein complex normally conducts calcium in primary cilia in response to fluid flow, and its dysfunction is thought to relieve calcium-dependent inhibition of adenylyl cyclases, raising cAMP levels, which are indeed elevated in kidney cysts in vivo. When the team treated dome tubuloids with 10 micromolar forskolin, the structures simply swelled — their area increased, consistent with chloride secretion into the existing lumen through the CFTR channel drawing in water osmotically. Suspension tubuloids did something far more interesting: they formed new cysts, with statistically significant increases in both cyst count and cyst area tracked over 72 hours, including live imaging every three hours in a system seeded with fluorescently labelled cells.

Confocal microscopy revealed the structural basis of this difference. Dome tubuloids display basal-out polarity: their primary cilia project from the apical membrane into the internal lumen, tight junctions marked by ZO-1 line the inner surface, and the basolateral membrane faces the collagen scaffold. Suspension tubuloids invert this arrangement entirely — an apical-out polarity, with cilia facing the outside and ZO-1 on the outer surface, consistent with previous reports that removing the extracellular matrix flips tubuloid polarity. Forskolin-treated suspension tubuloids formed cysts while retaining this inverted orientation, and the epithelial wall thinned significantly around the cystic regions, an observation the authors note has not previously been reported for this system. Dome tubuloids also showed significant wall thinning after treatment, along with dilation of their existing cystic structure.

One curious detail emerged from quantifying cilia: primary cilia on suspension tubuloids measured roughly 4 to 6 micrometres in length, longer than the 1 to 3 micrometres typical of healthy adult human kidney cilia, and longer than those on dome tubuloids. The authors suggest this elongation may reflect not only the polarity reversal but also the absence of fluid shear stress in static culture, though they flag the mechanism as a question for future work. The suspension cysts also expanded and constricted over time, breathing-like movements previously described in cysts from pluripotent stem cell-derived kidney organoids and attributed to epithelial rupture under expansive fluid pressure. Because the cysts formed within just three to five hours of treatment, the researchers argue that cAMP-driven proliferation alone cannot explain them, and point to fluid secretion through reoriented ion transporters such as CFTR and the sodium-potassium pump as a likely contributor that remains to be confirmed.

The study also demonstrated that both tubuloid types can be genetically modified via recombinant adeno-associated virus. Transduction with rAAV2/8 carrying a CMV-driven EGFP reporter produced fluorescent protein expression in both dome and suspension cultures, opening the door to introducing ciliary-localized optogenetic tools that could manipulate ciliary cAMP levels with light — an approach the same group has previously used to identify a cAMP signalosome in kidney epithelial cilia that drives cystogenesis through protein kinase A and the transcription factor CREB.

The authors are candid about limitations. Suspension tubuloids proliferate slowly and could not be expanded long-term, so the practical workflow derives them fresh from dome cultures at each passage; they also lack uniform size and structure, though microwell-based non-adherent platforms might standardize them. Dome tubuloids, meanwhile, can be maintained for around 90 days and at least five passages, while suspension cultures survived to about passage five, or roughly 50 days. Even so, the pair of systems now offers complementary views of the disease: dome tubuloids as a model of cyst enlargement in a regenerative, injury-repair-like epithelium, and suspension tubuloids as a mature epithelium capable of rapid, quantifiable, newly-formed cystogenesis. Together, the researchers conclude, they provide platforms for uncovering pathogenetic mechanisms underlying ADPKD and for testing candidate drugs in tissue-like, physiologically relevant microenvironments built from actual adult human kidney cells.

Subject of Research: Human kidney tubuloid cultures engineered to model cyst formation in autosomal dominant polycystic kidney disease

Article Title: Modelling kidney cystogenesis using human kidney tubuloid cultures

Article References: Rohe, J., Ashraf, A. P. K., Kehl, M., Thielmann, X., Ellinger, J., Kristiansen, G., Wachten, D., & Toma, M. I. (2026). Modelling kidney cystogenesis using human kidney tubuloid cultures. BMC Molecular and Cell Biology, 27(1), Article 26. https://doi.org/10.1186/s12860-026-00591-x

Image Credits: AI Generated

DOI: 10.1186/s12860-026-00591-x

Keywords: kidney organoids, tubuloids, polycystic kidney disease, ADPKD, cAMP signalling, cystogenesis, primary cilia, 3D cell culture, collecting duct, forskolin, rAAV transduction, epithelial polarity