Efficient and Rapid Induction of Human iPSCs/ESCs into Nephrogenic Intermediate Mesoderm Using Small Molecule-Based Differentiation Methods

Abstract

The first step in developing regenerative medicine approaches to treat renal diseases using pluripotent stem cells must be the generation of intermediate mesoderm (IM), an embryonic germ layer that gives rise to kidneys. In order to achieve this goal, establishing an efficient, stable and low-cost method for differentiating IM cells using small molecules is required. In this study, we identified two retinoids, AM580 and TTNPB, as potent IM inducers by high-throughput chemical screening, and established rapid (five days) and efficient (80% induction rate) IM differentiation from human iPSCs using only two small molecules: a Wnt pathway activator, CHIR99021, combined with either AM580 or TTNPB. The resulting human IM cells showed the ability to differentiate into multiple cell types that constitute adult kidneys, and to form renal tubule-like structures. These small molecule differentiation methods can bypass the mesendoderm step, directly inducing IM cells by activating Wnt, retinoic acid (RA), and bone morphogenetic protein (BMP) pathways. Such methods are powerful tools for studying kidney development and may potentially provide cell sources to generate renal lineage cells for regenerative therapy.

Cell Therapy Using Human Induced Pluripotent Stem Cell-Derived Renal Progenitors Ameliorates Acute Kidney Injury in Mice

“…We previously reported that retinoic acid receptor (RAR) agonists efficiently differentiated hiPSCs into IM cells [7]. Thus, the a-RAR agonist, TTNPB, was added to the EB method, which more efficiently generated OSR1 + cells (data not shown).…”

Novel regenerative therapy for acute kidney injury

“…Lineage-tracing analyses showed that Odd-skipped related 1 (Osr1) is one of the earliest markers of IM [44]. We generated OSR1-GFP knockin human iPSC lines and established efficient methods to induce OSR1 + IM cells using a combination treatment with growth factors and small molecules [45,46]. Osr1 is expressed in renal progenitors in IM and MM, although it is also expressed in the lateral plate mesoderm of early-stage mouse, chick, and fish embryos [44,47,48].…”

“…Our group has reported methods for inducing renal progenitor cells from human iPSCs/ESCs using stepwise differentiation and chemical biology strategies [41,45,46]. By screening growth factors and chemical compounds that can induce renal progenitor cells from human iPSCs, we could induce OSR1 + IM cells and OSR1 + SIX2 + renal progenitors.…”

“…By screening growth factors and chemical compounds that can induce renal progenitor cells from human iPSCs, we could induce OSR1 + IM cells and OSR1 + SIX2 + renal progenitors. We also reconstituted 3D nephron-like structures in vitro that mainly contain proximal renal tubules [41,45,46]. Xia et al reported a protocol by which human iPSCs/ ESCs were differentiated into the progenitors like ureteric bud (UB), an embryonic cell population that gives rise to the collecting system and the lower urinary tract from the renal pelvis to a part of the urinary bladder [52].…”

Human iPSC Models: A Platform for Investigating Neurodevelopmental Diseases

“…With this innovative methodology, many laboratories are using hiPSC to model human diseases that occur in many organ systems [6][7][8][9][10][11][12][13][14]. While hiPSC are a promising investigative method to define molecular deficits in disease cells, it will require rigorous comparisons between many patients with the same disease to clearly characterize cellular dysfunction and abnormalities [15].…”