Cookies on this website
We use cookies to ensure that we give you the best experience on our website. If you click 'Continue' we'll assume that you are happy to receive all cookies and you won't see this message again. Click 'Find out more' for information on how to change your cookie settings.
Skip to main content

OBJECTIVE: To investigate the efficacy of CRISPR-Cas9 mediated editing in human chondrocytes, and to develop a genome editing approach relevant to cell-based repair. METHODS: Transfection of human articular chondrocytes (both healthy and osteoarthritic) with ribonucleoprotein complexes (RNP) containing Cas9 and a crisprRNA targeting exon2 of MMP13 was performed to assess editing efficiency and effects on MMP13 protein levels and enzymatic activity. Using spheroid cultures, protein levels of a major target of MMP13, type II collagen, were assessed by western blot and immunofluorescence. RESULTS: With an editing efficiency of 63-74%, secreted MMP13 protein levels and activity were significantly reduced (percentage decrease 34.14% without and 67.97% with IL-1β based on median values of MMP13 enzymatic activity, N = 7) comparing non-edited with edited cell populations using an exon-targeting gRNA resulting in premature stop codons through non-homologous end joining (NHEJ). Accumulation of cartilage matrix protein type II collagen was enhanced in edited cells in spheroid culture, compared to non-edited controls. CONCLUSION: CRISPR-Cas9 mediated genome editing can be used to efficiently and reproducibly establish populations of human chondrocytes with stably reduced expression of key genes of interest without the need for clonal selection. Such an editing approach has the potential to greatly enhance current cell-based therapies for cartilage repair.

Original publication

DOI

10.1016/j.joca.2018.09.001

Type

Journal article

Journal

Osteoarthritis cartilage

Publication Date

01/2019

Volume

27

Pages

140 - 147

Keywords

3D spheroids, Autologous chondrocyte implantation, CRISPR-Cas9, Cartilage, MMP13