Increased TGF-β and BMP levels and improved chondrocyte-specific marker expression in vitro under cartilage-specific physiological osmolarity

Ufuk Tan Timur*, Marjolein Caron, Guus van den Akker, Anna van der Windt, Jenny Visser, Lodewijk van Rhijn, Harrie Weinans, Tim Welting, Pieter Emans, Holger Jahr

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

22 Citations (Scopus)
4 Downloads (Pure)


During standard expansion culture (i.e., plasma osmolarity, 280 mOsm) human articular chondrocytes dedifferentiate, making them inappropriate for autologous chondrocyte implantation to treat cartilage defects. Increasing the osmolarity of culture media to physiological osmolarity levels of cartilage (i.e., 380 mOsm), increases collagen type II (COL2A1) expression of human articular chondrocytes in vitro, but the underlying molecular mechanism is not fully understood. We hypothesized that TGF-β superfamily signaling may drive expression of COL2A1 under physiological osmolarity culture conditions. Human articular chondrocytes were cultured in cytokine-free medium of 280 or 380 mOsm with or without siRNA mediated TGF-β2 knockdown (RNAi). Expression of TGF-β isoforms, and collagen type II was evaluated by RT-qPCR and immunoblotting. TGF-β2 protein secretion was evaluated using ELISA and TGF-β bioactivity was determined using an established reporter assay. Involvement of BMP signaling was investigated by culturing human articular chondrocytes in the presence or absence of BMP inhibitor dorsomorphin and BMP bioactivity was determined using an established reporter assay. Physiological cartilage osmolarity (i.e., physosmolarity) most prominently increased TGF-β2 mRNA expression and protein secretion as well as TGF-β bioactivity. Upon TGF-β2 isoform-specific knockdown, gene expression of chondrocyte marker COL2A1 was induced. TGF-β2 RNAi under physosmolarity enhanced TGF-β bioactivity. BMP bioactivity increased upon physosmotic treatment, but was not related to TGF-β2 RNAi. In contrast, dorsomorphin inhibited COL2A1 mRNA expression in human articular chondrocytes independent of the osmotic condition. Our data suggest a role for TGF-β superfamily member signaling in physosmolarity-induced mRNA expression of collagen type II. As physosmotic conditions favor the expression of COL2A1 independent of our manipulations, contribution of other metabolic, post-transcriptional or epigenetic factors cannot be excluded in the underlying complex and interdependent regulation of marker gene expression. Dissecting these molecular mechanisms holds potential to further improve future cell-based chondral repair strategies.

Original languageEnglish
Article number795
JournalInternational Journal of Molecular Sciences
Issue number4
Publication statusPublished - 13 Feb 2019

Bibliographical note

Funding Information:
Funding: H.J.: H.W. and U.T.T. are members of the D-BOARD Consortium; This project has received funding from the European Union’s Seventh Framework Programme for Research, Technological Development, and Demonstration under Grant Agreement no. 305815 (HEALTH.2012.2.4.5-2). We are further indebted to START of the Faculty of Medicine of the RWTH Aachen and the Aachen-Jülich-Haifa Umbrella Cooperation (acronym SmartCart). This project was also financially supported by the Dutch Arthritis Foundation (grant LLP14).

Publisher Copyright:
© 2019 by the authors. Licensee MDPI, Basel, Switzerland.


Dive into the research topics of 'Increased TGF-β and BMP levels and improved chondrocyte-specific marker expression in vitro under cartilage-specific physiological osmolarity'. Together they form a unique fingerprint.

Cite this