Differentiation of Hypertrophic Chondrocytes from Human iPSCs for the In Vitro Modeling of Chondrodysplasias

Abstract

Summary Chondrodysplasias are hereditary diseases caused by mutations in the components of growth cartilage. Although the unfolded protein response (UPR) has been identified as a key disease mechanism in mouse models, no suitable in vitro system has been reported to analyze the pathology in humans. Here, we developed a three-dimensional culture protocol to differentiate hypertrophic chondrocytes from induced pluripotent stem cells (iPSCs) and examine the phenotype caused by MATN3 and COL10A1 mutations. Intracellular MATN3 or COL10 retention resulted in increased ER stress markers and ER size in most mutants, but activation of the UPR was dependent on the mutation. Transcriptome analysis confirmed a UPR with wide-ranging changes in bone homeostasis, extracellular matrix composition, and lipid metabolism in the MATN3 T120M mutant, which further showed altered cellular morphology in iPSC-derived growth-plate-like structures in vivo . We then applied our in vitro model to drug testing, whereby trimethylamine N-oxide led to a reduction of ER stress and intracellular MATN3.

The Induced Pluripotent Stem Cells in Articular Cartilage Regeneration and Disease Modelling: Are We Ready for Their Clinical Use?

“…Another group of disorders related to chondrogenesis that was studied using iPSC modelling is connected with the accumulation of proteins in the intracellular compartments of chondrocytes, leading to the activation of the unfolded protein response (UPR) [171,172,182]. Among them, we may distinguish multiple epiphyseal dysplasia (MED) and metaphyseal chondrodysplasia type Schmid (MCDS), which are related to the widening and irregularity of growth plates, an increased risk of developing early osteoarthritis of the knee and hip [171,183,184].…”

“…Another group of disorders related to chondrogenesis that was studied using iPSC modelling is connected with the accumulation of proteins in the intracellular compartments of chondrocytes, leading to the activation of the unfolded protein response (UPR) [171,172,182]. Among them, we may distinguish multiple epiphyseal dysplasia (MED) and metaphyseal chondrodysplasia type Schmid (MCDS), which are related to the widening and irregularity of growth plates, an increased risk of developing early osteoarthritis of the knee and hip [171,183,184]. These diseases are related to the occurrence of mutations in type 10 collagen (COL10A1), a marker of hypertrophic chondrocytes, and matrilin-3 (MATN3), the protein responsible for terminal chondrogenic differentiation and homeostasis of the cartilage [171,181,185,186].…”

“…Among them, we may distinguish multiple epiphyseal dysplasia (MED) and metaphyseal chondrodysplasia type Schmid (MCDS), which are related to the widening and irregularity of growth plates, an increased risk of developing early osteoarthritis of the knee and hip [171,183,184]. These diseases are related to the occurrence of mutations in type 10 collagen (COL10A1), a marker of hypertrophic chondrocytes, and matrilin-3 (MATN3), the protein responsible for terminal chondrogenic differentiation and homeostasis of the cartilage [171,181,185,186]. A recently published model of this disease by the Pretemer team revealed that the endoplasmic reticulum was significantly enlarged after the chondrogenic differentiation of COL10A1 and MATN3 mutants [171].…”

“…These diseases are related to the occurrence of mutations in type 10 collagen (COL10A1), a marker of hypertrophic chondrocytes, and matrilin-3 (MATN3), the protein responsible for terminal chondrogenic differentiation and homeostasis of the cartilage [171,181,185,186]. A recently published model of this disease by the Pretemer team revealed that the endoplasmic reticulum was significantly enlarged after the chondrogenic differentiation of COL10A1 and MATN3 mutants [171]. Histological staining also revealed the increased deposition of COL10 and matrilin-3 intracellularly in several mutants, a state partially reversed by using trimethylamine N-oxide (TMAO), an agent acting as a chemical chaperone [171].…”

“Lessons from Rare Forms of Osteoarthritis”

“…The inhibition of the pathways that these proteins are involved in alleviates chondrodysplasia by preventing aberrant chondrocyte differentiation [72,73]. Positive reduction in ER stress has been shown to occur in vitro when cells are treated with trimethylamine N-oxide as well as lesser accumulation of defective protein in cells [74,75].…”