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Method for differentiating induced pluripotent stem cells into renal proximal tubular cell-like cells

A technology of pluripotent stem cells and renal proximal tubules, applied in the field of generating renal proximal tubule cell-like cells, which can solve the problems of limited application

Active Publication Date: 2016-10-12
AGENCY FOR SCI TECH & RES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Further, hESC-derived cells express some major drug transporters, such as organic anion transporter (OAT) 3, at very low levels, which may limit the application of such cells in in vitro renal toxicology

Method used

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  • Method for differentiating induced pluripotent stem cells into renal proximal tubular cell-like cells
  • Method for differentiating induced pluripotent stem cells into renal proximal tubular cell-like cells
  • Method for differentiating induced pluripotent stem cells into renal proximal tubular cell-like cells

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Experimental program
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Effect test

Embodiment 1

[0160] Beginning on November 5, 2012, hiPSC-derived HTPC-like cells were treated with two test drugs (rifampicin and paraquat, both nephrotoxicants directly toxic to PTC). Drug response was tested using an in vitro model of renal toxicology based on interleukin 6 / interleukin 8 expression (Li et al., Toxicol. Res. 2 (2013), 352-362). The data show that IL-6 (and in the case of rifampin, IL-8) expression is increased after treatment with PTC-specific nephrotoxicants. This suggests that treatment with PTC-specific nephrotoxicants resulted in a positive response.

[0161] Materials and methods

[0162] All steps are performed using aseptic technique.

[0163] Reagents: mTeSR1 medium Kit: STEM CELL Technologies, Cat. No.: 05850; BD Matrigel TM Basement Membrane Matrix, Low Growth Factor (GFR), BD Biosciences, Cat. No. 356230; Rock Inhibitor (Y-27632) Calbiochem, Cat. No. 688000; Renal Epithelial Basal Medium (REBM), Lonza Biosciences, Cat. No. CC-3191; Cell Isolation Reagent, ...

Embodiment 2

[0189] Using a one-step method for differentiating hiPSCs into HPTC-like cells, drug testing was performed immediately without prior cell collection. The predictive performance of hiPSC-derived cells was determined by evaluating the response to 30 compounds. Automatic classification of results by support vector machine algorithm. Using this method, PTC toxicity can be predicted with high accuracy in humans. Therefore, the one-step differentiation approach combined with machine learning may provide an efficient method for accurately predicting PTC toxicity in humans.

[0190] Materials and methods

[0191] Expansion and differentiation of hiPSCs: iPS(foreskin)-4, iPSIMR90-4 and iPS DF19-9-11T.H cells were obtained from WiCell Research Institute (Madison, WI, USA). Work with these hiPSC lines was approved by the Institutional Review Board of the National University of Singapore (NUS-IRB reference code: 13-437). Undifferentiated cells were expanded in multi-well plates coated...

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Abstract

There is provided a method of differentiating an induced pluripotent stem cell (iPSC) into a renal proximal tubular cell (PTC)-like cell. The method comprises culturing an undifferentiated iPSC in a renal epithelial cell culture medium in the presence of one or more extracellular matrix (ECM) molecules, bone morphogenic protein 2 (BMP2) and bone morphogenic protein 7 (BMP7), for a period of from about 8 to about 10 days, under conditions sufficient to induce differentiation of the iPSC into a PTC-like cell. A cell population of differentiated PTC-like cells is also provided, as well as uses and methods of use of the cell population.

Description

[0001] Cross References to Related Applications [0002] This application claims the benefit and priority of Singapore Provisional Application No. 201308341-5 filed on 11.11.2013, the contents of which are hereby incorporated herein by reference. technical field [0003] The present invention relates to a method for producing renal proximal tubular cell-like cells by differentiation of induced pluripotent stem cells. Background technique [0004] Renal epithelial cells are useful in treating kidney disease. This includes applications in bioartificial kidney devices to replace or compensate for lost organ function. Therefore, an adequate supply of renal epithelial cells is required. Of particular interest are renal proximal tubular cells (PTCs), which perform a variety of renal functions. [0005] Renal proximal tubular cells play a role in drug transport and metabolism and are therefore also of particular interest in in vitro renal toxicology. This cell type is the main ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071A61K35/545
CPCA61K35/545C12N5/0686C12N2501/155C12N2501/727C12N2506/45C12N2533/90A61K35/22A61L27/3604A61L27/3641A61L27/3839A61L27/3895A61L27/54A61L27/58A61L2300/414A61L2300/64A61L2430/26A61L2430/40
Inventor D·津克应仪如
Owner AGENCY FOR SCI TECH & RES
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