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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 (ROKβ) and ROCK2 (ROKα), 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 αSMA expression. Furthermore, our data confirm that the inhibition of ROCK signalling by a pharmacological inhibitor (i.e. Y27632) attenuates stiffness induced αSMA 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 αSMA expression in myofibroblasts on stiff matrix as opposed to soft matrix where αSMA expression was reduced significantly. Paradoxically, on stiff matrix, the absence of one isoform (particularly ROCK2) exaggerated αSMA expression and led to thick fibre assembly. Moreover complete loss of αSMA 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.

Journal Article Type Article
Publication Date 2017-06
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
APA6 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
DOI https://doi.org/10.1165/rcmb.2016-0306OC
Publisher URL http://www.atsjournals.org/doi/10.1165/rcmb.2016-0306OC
Copyright Statement Copyright information regarding this work can be found at the following address: http://eprints.nottingh.../end_user_agreement.pdf

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Copyright Statement
Copyright information regarding this work can be found at the following address: http://eprints.nottingham.ac.uk/end_user_agreement.pdf





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