Use of Cartilage Derived From Murine Induced Pluripotent Stem Cells for Osteoarthritis Drug Screening

Abstract

Objective The discovery of novel disease-modifying drugs for osteoarthritis (OA) is limited by the lack of adequate genetically-defined cartilage tissues for application in high-throughput screening systems. We addressed this need by synthesizing cartilage from induced pluripotent stem cells (iPSCs) to establish and validate an in vitro model of OA. Methods iPSC-derived or native mouse cartilage samples were treated with the cytokine interleukin-1α (IL-1α) for 3 days to model the inflammatory environment of OA. Biochemical content, mechanical properties, and gene expression of the resulting tissues were assayed. In addition, the inflammatory and catabolic environment of the media was assessed. To establish high-throughput capability, we utilized a 96-well plate format and conducted a screen of previously identified candidate OA drugs. Glycosaminoglycan release into the media was used as the primary output for screening. Results Treatment of iPSC-derived or native cartilage with IL-1α induced characteristic features of OA in a rapid and dose-dependent manner. In addition to the loss of glycosaminoglycans and tissue mechanical properties, IL-1α treatment induced expression of matrix metalloproteinases and increased production of the inflammatory mediators nitric oxide and prostaglandin E2. In the high-throughput screen validation, all candidate OA therapeutics provided some benefit, but only the NF-κB inhibitor SC-514 effectively reduced cartilage loss in response to IL-1α. Conclusions This work demonstrates the utility of iPSCs for studying cartilage pathology, and provides a platform for identifying novel, patient-specific therapeutics that prevent cartilage degradation and modify the course of OA development.

CRISPR/Cas9 Editing of Murine Induced Pluripotent Stem Cells for Engineering Inflammation‐Resistant Tissues

“…No significant effects of IL-1α were found for total collagen or total collagen/DNA (Fig. S1B,C), consistent with previous studies (26) and possibly due to the retention of partially degraded, large macromolecular constituents from enzymatically degraded tissue (18). …”

“…Importantly, cartilage derived from CRISPR/Cas9-edited iPSCs displayed the capacity to withstand treatment with 1 ng/ml IL-1α. Consistent with our work, previous studies have shown that treatment with 1 ng/ml IL-1α results in severe degradation of both engineered cartilage derived from Il1r1-expressing cells as well as cartilage from mouse explants (17, 18, 26). However, in our study, engineered tissue derived from Il1r1 -/- cells was protected from an in vitro model of OA.…”

TRPV4 as a therapeutic target for joint diseases

“…In particular, iPSCs can be generated from patients with TRPV4 mutations that cause the spectrum of skeletal dysplasias (Saitta et al, 2014) and the arthopathies, or from patients with osteoporosis, rheumatoid arthritis, or osteoarthritis. Cartilage derived from iPSCs has been shown to replicate various aspects of human disease in vitro (Willard et al, 2014) and can be used to test therapeutic approaches for the spectrum of joint diseases.…”

Genome Engineering for Personalized Arthritis Therapeutics

“…Nf-κB). [100] Such findings demonstrate the use of an in vitro arthritis disease modeling platform to identify candidate DMOADs. Future efforts should consider extending this approach to human iPSC-based OA modeling platforms, where human iPSC-derived cartilage tissue might be exposed to OA-relevant stimuli, such as inflammatory cytokines and/or mechanical loading (Key figure, Figure 1).…”

“…Recent efforts directed towards arthritis disease modeling have shown that administration of inflammatory cytokines IL1 and TNF to engineered tissues derived from murine iPSCs can result in tissue degradation, which is consistent with the detrimental effects of these inflammatory cytokines in the joints of patients harboring various forms of arthritis. [95,100] Degradative changes in induced cartilage might thus be ameliorated through small molecule-mediated inhibition of catabolic enzymes (e.g. Mmp-13) or transcription factors (e.g.…”