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Role of ROCK isoforms in regulation of stiffness induced myofibroblast differentiation in lung fibrosis

Htwe, Su S.; Cha, Byung H.; Yue, Kan; Khademhosseini, Ali; Knox, Alan J.; Ghaemmaghami, Amir M.

Authors

Su S. Htwe

Byung H. Cha

Kan Yue

Ali Khademhosseini

Alan J. Knox



Abstract

Fibrosis is a major cause of progressive organ dysfunction in several chronic pulmonary diseases. Rho associated coiled-coil forming kinase (ROCK) has shown to be involved in myofibroblast differentiation driven by altered matrix stiffness in fibrotic state. There are two known ROCK isoforms in human, ROCK1 (ROK?) and ROCK2 (ROK?), but specific role of each isoform in myofibroblast differentiation in lung fibrosis remains unknown. To study this, we developed a Gelatin methacryloyl (GelMA) hydrogel based culture system with different stiffness levels relevant to healthy and fibrotic lungs. We have shown that stiff matrix and not soft matrix, can induce myofibroblast differentiation with high ?SMA expression. Furthermore, our data confirm that the inhibition of ROCK signalling by a pharmacological inhibitor (i.e. Y27632) attenuates stiffness induced ?SMA expression and fibre assembly in myofibroblasts. To assess the role of ROCK isoforms in this process we used siRNA to knock down the expression of each isoform. Our data showed that knocking down either ROCK1 or ROCK2 did not result in a reduction in ?SMA expression in myofibroblasts on stiff matrix as opposed to soft matrix where ?SMA expression was reduced significantly. Paradoxically, on stiff matrix, the absence of one isoform (particularly ROCK2) exaggerated ?SMA expression and led to thick fibre assembly. Moreover complete loss of ?SMA fibre assembly was seen only in the absence of both ROCK isoforms suggesting that both isoforms are implicated in this process. Overall our results indicate the differential role of ROCK isoforms in myofibroblast differentiation on soft and stiff matrices.

Citation

Htwe, S. S., Cha, B. H., Yue, K., Khademhosseini, A., Knox, A. J., & Ghaemmaghami, A. M. (2017). Role of ROCK isoforms in regulation of stiffness induced myofibroblast differentiation in lung fibrosis. American Journal of Respiratory Cell and Molecular Biology, 56(6), 772-783. https://doi.org/10.1165/rcmb.2016-0306OC

Journal Article Type Article
Acceptance Date Feb 17, 2017
Online Publication Date Feb 22, 2017
Publication Date 2017-06
Deposit Date Feb 28, 2017
Publicly Available Date Mar 28, 2024
Journal American Journal of Respiratory Cell and Molecular Biology
Print ISSN 1044-1549
Electronic ISSN 1535-4989
Publisher American Thoracic Society
Peer Reviewed Peer Reviewed
Volume 56
Issue 6
Pages 772-783
DOI https://doi.org/10.1165/rcmb.2016-0306OC
Public URL https://nottingham-repository.worktribe.com/output/863110
Publisher URL http://www.atsjournals.org/doi/10.1165/rcmb.2016-0306OC