Many diseases and drugs affect kidney function by damaging the glomeruli — the major site of blood filtration and a target for many diseases. An in vitro model of the human glomerulus could facilitate therapeutic discovery and illuminate kidney disease mechanisms. Efforts to develop such models have been limited by the lack of a reliable source of functional human podocytes, the specialized epithelial cells that regulate selective permeability in the kidney glomerulus. Despite recent advances in generating nephron progenitors and multicellular organoid structures, a method for the differentiation of human induced pluripotent stem (hiPS) cells into functional podocytes with specificity and efficiency remains elusive.

Fig. 1. Directed differentiation of human iPS cells into mature kidney cells. (A) Schematic illustration of the method for human iPS cell differentiation into kidney glomerular podocytes. (B) Transcriptome analysis of genes involved in different stages of kidney development. Red, up-regulated; Blue, down-regulated. (C) Immunofluorescence staining showing the expression of podocin (green), a podocyte lineage identification marker, in the human iPS cell-derived kidney cells. (D) Scanning electron micrographs of human iPS cell-derived kidney podocytes compared to an established human podocyte cell line (PCL) which lacks morphological features necessary for kidney function.

To address these challenges, we developed a highly efficient method for directed differentiation of hiPS cells into functional kidney glomerular podocytes which was enabled by the discovery of key signaling molecules that support adhesion and stimulate gene regulatory pathways in human kidney development. The resulting hiPS cell-derived podocytes express markers consistent with mature phenotype, exhibit primary and secondary foot processes, and integrate into forming glomerular structures when injected into mouse embryonic kidney explants.

We leveraged the hiPS cell-derived podocytes and organs-on-chips technology to engineer an in vitro model of a functional human kidney glomerular capillary wall. Unlike previously reported kidney organoids that lack fluid path or circuitry and have limited control over tissue structure and function, we found that our human glomerulus-on-a-chip supports podocyte differentiation and recapitulates the tissue-tissue interface (podocytes―basement membrane―endothelium) and selective molecular filtration function of the glomerulus. Our results demonstrate the feasibility of generating mature podocytes from unspecialized stem cells in a robust manner, providing a unique opportunity to engineer a functional human kidney glomerulus-on-a-chip. These results could facilitate investigations that aim to uncover the molecular and cellular mechanisms of human kidney development, function, and disease progression.

Fig. 2. Establishment of a functional human kidney glomerulus-on-a-chip. (A) Microfluidic organ-on-a-chip device showing dedicated compartments to mimic the vasculature (capillary channel) and podocyte layer (urinary channel). (B) 3D reconstruction of the human kidney’s blood filtration barrier modeled using iPS cell-derived podocytes (green) and primary glomerular endothelium (magenta). (C) Selective filtration of molecules in the engineered glomerulus-on-a-chip compared to an estimated in vivo values. (D) Modeling podocyte injury induced by the chemotherapy drug Adriamycin. Scanning electron micrographs showing damage to podocytes exposed to clinically relevant concentration of the drug. (E) Glomerulus-on-a-chip models adriamycin-induced albuminuria. Bar graphs represent the concentration of albumin in the urinary compartment or loss of albumin protein into urine, a hallmark of kidney disease.

These results also provide a low-cost alternative to animal models for nephrotoxicity testing, drug screening, and the development of novel therapeutic strategies for both genetic and acquired forms of human kidney diseases. Given that specialized podocytes in a fully formed human kidney are post-mitotic and unable to undergo regenerative proliferation to compensate for cell loss or an increase in glomerular basement membrane surface area in vivo, our results also provide opportunities for cell-based therapy and regenerative medicine approaches for kidney disease, including chronic kidney disease and podocytopathies.

Samira Musah
Department of Biomedical Engineering, Pratt School of Engineering, Duke University, Durham, NC, USA
Department of Medicine, Division of Nephrology, Duke University School of Medicine, Durham, NC, USA

Publications

Directed differentiation of human induced pluripotent stem cells into mature kidney podocytes and establishment of a Glomerulus Chip.
Samira Musah, Nikolaos Dimitrakakis, Diogo M. Camacho, George M. Church, Donald E. Ingber
Nat Protoc. 2018 Jul

Mature induced-pluripotent-stem-cell-derived human podocytes reconstitute kidney glomerular-capillary-wall function on a chip.
Musah S, Mammoto A, Ferrante TC, Jeanty SSF, Hirano-Kobayashi M, Mammoto T, Roberts K, Chung S, Novak R, Ingram M, Fatanat-Didar T, Koshy S, Weaver JC, Church GM, Ingber DE
Nat Biomed Eng. 2017

Read offline:     

Related Articles:

	Molecular classification of glioma stem cells Comprehensive genetic analysis of glioblastomas by The Cancer Genome Atlas (TCGA) has proposed a new classification based on genetic abnormalities. The characteristic genetic abnormalities identified in the TCGA are promising…
	The algal organic matter: A novel carbon source to… In recent years, nitrogen pollution, especially nitrate is an important problem faced by lots of reservoirs. Excessive nitrogen levels can cause water eutrophication, resulting in deterioration of water quality by…
	Rabbits with mammary carcinomas as a model for… Within a breeding colony of rabbits, the American pathologist Harry Greene (1904-1969) observed that mammary carcinomas were restricted to certain families. This is suggestive of a familiar predisposition as it…
	Gas types affect stability of microbubbles and… Ultrasound (US) is used for clinical diagnosis and therapy. Microbubbles (MBs) as US-contrast agents are applied to detect micro blood flow or specific lesions. Recently, the combination of MBs and…
	Fast artificial intelligence for detecting the invisible Artificial Intelligence (AI) relies on synthetic neural networks, inspired by functional properties of living intelligent systems (brains) and learns autonomously on the basis of unsupervised learning procedures. Such learning permits,…
	Can RNA therapeutics help spinal cord injury treatments? In the central nervous system (CNS) nerve regeneration is severely hampered making recovery from lesions such as spinal cord injury (SCI) extremely difficult and one of the most formidable challenges…