Differentiation of Human Induced Pluripotent Stem Cells (iPSCs)–derived Mesenchymal Progenitors into Chondrocytes

• Type of Material: Article
• Authors: Nazir Khan, Emory University; Martha Elena Diaz, Emory University; Hicham Drissi, Emory University
• Language: English
• Date: 2023-11-05
• Publisher: Bio-Protocol LLC
• Publication Version: Final Published Version
• Copyright Statement: © 2023 The Authors; This is an open access article under the CC BY license
• License: Creative Commons Attribution 4.0 International
• Final Published Version (URL): https://doi.org/10.21769/BioProtoc.4874
• Title of Journal or Parent Work: Bio-Protocol
• Volume: 13
• Issue: 21
• Start Page: e4874
• Grant/Funding Information: This work was supported by funds from Veteran Affairs and Emory University School to Medicine. This research was funded by Georgia CTSA/REM Pilot Project 00080502 to H.D., Veteran Affairs CaReAP Award (I01-BX004878) to H.D.
• Keywords: High density culture; Chondrogenic differentiation; Chondrocytes; Induced pluripotent stem cells (iPSCs); Mesenchymal stromal/stem cells (MSCs)
• Research Categories: Biology, Cell; Engineering, Biomedical

Abstract

Induced pluripotent stem cells (iPSCs) generated from human sources are valuable tools for studying skeletal development and diseases, as well as for potential use in regenerative medicine for skeletal tissues such as articular cartilage. To successfully differentiate human iPSCs into functional chondrocytes, it is essential to establish efficient and reproducible strategies that closely mimic the physiological chondrogenic differentiation process. Here, we describe a simple and efficient protocol for differentiation of human iPSCs into chondrocytes via generation of an intermediate population of mesenchymal progenitors. These methodologies include step-by-step procedures for mesenchymal derivation, induction of chondrogenic differentiation, and evaluation of the chondrogenic marker gene expression. In this protocol, we describe the detailed procedure for successful derivation of mesenchymal progenitor population from human iPSCs, which are then differentiated into chondrocytes using high-density culture conditions by stimulating with bone morphogenetic protein-2 (BMP-2) or transforming growth factor beta-3 (TGFβ-3). The differentiated iPSCs exhibit temporal expression of cartilage genes and accumulation of a cartilaginous extracellular matrix in vitro, indicating successful chondrogenic differentiation. These detailed methodologies help effective differentiation of human iPSCs into the chondrogenic lineage to obtain functional chondrocytes, which hold great promise for modeling skeletal development and disease, as well as for potential use in regenerative medicine for cell-based therapy for cartilage regeneration.