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Articular cartilage has a poor capacity for self-repair, consequently, defects are one of the major causes of immobility and poor quality of life for millions of individuals worldwide. Damage to articular cartilage can be caused by trauma, ageing, genetic factors and degenerative diseases such as osteoarthritis and is often irreversible without surgical intervention. The current clinically approved therapies for cartilage repair rely on the use of autologous chondrocytes and can be limited by chondrocyte expansion and generation of fibrous repair tissues with inferior mechanical properties. This has led to the emergence of alternative cartilage tissue engineering (CTE) as a strategy to regenerate long-lasting functional tissues.
Human pluripotent stem cells (hPSCs) show exciting potential as an alternative cell source for cartilage regeneration. This is due to their capacity for unlimited self-renewal and ability to differentiate into almost all cell types within the body, thus forming an unlimited supply of cells for regenerative therapies. Signalling from the TGF-β growth factor family plays a critical role in regulating chondrogenic differentiation and maintenance of articular cartilage tissues. However, growth factor-driven chondrogenic differentiation is limited by the intrinsic properties of growth factor peptides such as short half-life, rapid denaturation in vivo and high costs[1].
Graphene oxide (GO) is a 2D carbon nanomaterial that has gained attention worldwide due to its extraordinary physiochemical properties. The advantage of GO over other nanomaterials is its ability to act as a multifunctional bioactive moiety, stemming from its unique 2D structure and rich surface chemistry which facilitates functionalisation. GO has been shown to improve chondrogenesis following the absorption of proteins and growth factors onto the surface of GO flakes or through improving mechanical properties within hydrogel scaffolds. However few studies have investigated the intrinsic chondroinductive effects of GO alone.
Here we demonstrate that GO alone is able to stimulate the TGF-β signalling pathway in human chondrocytes (TC28a2) and hPSC derived chondroprogenitors, thus highlighting a potentially new and cost-efficient means of inducing cartilage regeneration. Induction of the canonical TGFB signalling pathway was demonstrated through the use of a TGFβ signalling reporter (SBE-nLUCp) [2] and further validated by pSMAD2 immunoblotting and analysis of TGFB response genes via RT-QPCR. Exploiting the unique intrinsic fluorescence of our GO enabled robust analysis of interactions between GO and single chondrocytes through confocal live-cell imaging using the method developed by Vranic et al [3]. This facilitated an in-depth understanding of the mechanisms by which GO increases TGFB signalling activity.
[1] M. J. S. Ferreira et al., “Pluripotent stem cells for skeletal tissue engineering,”, 2021, doi: 10.1080/07388551.2021.1968785.
[2] S. Woods et al., “Regulation of TGFbeta Signalling by TRPV4 in Chondrocytes,” Cells, vol. 10, no. 4, 2021, doi: 10.3390/cells10040726.
[3] S. Vranic et al., “Live Imaging of Label-Free Graphene Oxide Reveals Critical Factors Causing Oxidative-Stress-Mediated Cellular Responses,” ACS Nano, vol. 12, no. 2, pp. 1373–1389, 2018, doi: 10.1021/acsnano.7b07734.
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