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      Photoelasticity-based evaluation of cellular contractile force for phenotypic discrimination of vascular smooth muscle cells

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          Abstract

          Vascular smooth muscle cells (VSMCs) have two distinct phenotypes: contractile and synthetic. The major difference between these phenotypes lies in the magnitude of the contractile force produced by the cell. Although traction force microscopy (TFM) is often used to evaluate cellular contractile force, this method requires complex preprocessing and a sufficiently compliant substrate. To evaluate the contractile force and the phenotype of living VSMCs with minimal effort and in a manner independent of the substrate stiffness, we propose a photoelasticity-based method using retardation, which is related to the difference between the first and second principal stresses and their orientation. The results demonstrate that actin filaments co-localize with areas of high retardation in cells, indicating that the retardation of VSMCs is promoted by actin filaments. The retardation of cells treated with calyculin A and Y-27632 tended to be larger and smaller, respectively, than that of control cells. Cell traction force significantly correlates with total cell retardation ( r 2 = 0.38). The retardation of contractile VSMCs (passage 2) was significantly higher than that of synthetic VSMCs (passage 12). These results indicate that cell retardation can be used to assess cell contractile force and, thus, determine the phenotype of VSMCs.

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          The effect of matrix stiffness on the differentiation of mesenchymal stem cells in response to TGF-β.

          Bone marrow mesenchymal stem cells (MSCs) are a valuable cell source for tissue engineering and regenerative medicine. Transforming growth factor β (TGF-β) can promote MSC differentiation into either smooth muscle cells (SMCs) or chondrogenic cells. Here we showed that the stiffness of cell adhesion substrates modulated these differential effects. MSCs on soft substrates had less spreading, fewer stress fibers and lower proliferation rate than MSCs on stiff substrates. MSCs on stiff substrates had higher expression of SMC markers α-actin and calponin-1; in contrast, MSCs on soft substrates had a higher expression of chondrogenic marker collagen-II and adipogenic marker lipoprotein lipase (LPL). TGF-β increased SMC marker expression on stiff substrates. However, TGF-β increased chondrogenic marker expression and suppressed adipogenic marker expression on soft substrates, while adipogenic medium and soft substrates induced adipogenic differentiation effectively. Rho GTPase was involved in the expression of all aforementioned lineage markers, but did not account for the differential effects of substrate stiffness. In addition, soft substrates did not significantly affect Rho activity, but inhibited Rho-induced stress fiber formation and α-actin assembly. Further analysis showed that MSCs on soft substrates had weaker cell adhesion, and that the suppression of cell adhesion strength mimicked the effects of soft substrates on the lineage marker expression. These results provide insights of how substrate stiffness differentially regulates stem cell differentiation, and have significant implications for the design of biomaterials with appropriate mechanical property for tissue regeneration. Copyright © 2011 Elsevier Ltd. All rights reserved.
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            Regulation and characteristics of vascular smooth muscle cell phenotypic diversity

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              Silicone rubber substrata: a new wrinkle in the study of cell locomotion.

              When tissue cells are cultured on very thin sheets of cross-linked silicone fluid, the traction forces the cells exert are made visible as elastic distortion and wrinkling of this substratum. Around explants this pattern of wrinkling closely resembles the "center effects" long observed in plasma clots and traditionally attributed to dehydration shrinkage.
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                Author and article information

                Contributors
                sugita.shukei@nitech.ac.jp
                Journal
                Sci Rep
                Sci Rep
                Scientific Reports
                Nature Publishing Group UK (London )
                2045-2322
                8 March 2019
                8 March 2019
                2019
                : 9
                : 3960
                Affiliations
                [1 ]ISNI 0000 0001 0656 7591, GRID grid.47716.33, Department of Electrical and Mechanical Engineering, , Graduate School of Engineering, Nagoya Institute of Technology, ; Nagoya, Japan
                [2 ]ISNI 0000 0001 0656 7591, GRID grid.47716.33, Department of Techno-Business Administration, , Graduate School of Engineering, Nagoya Institute of Technology, ; Nagoya, Japan
                [3 ]ISNI 0000 0001 0943 978X, GRID grid.27476.30, Department of Mechanical Systems Engineering, , Graduate School of Engineering, Nagoya University (Present), ; Nagoya, Japan
                Author information
                http://orcid.org/0000-0002-0922-2465
                Article
                40578
                10.1038/s41598-019-40578-7
                6408479
                30850684
                a2e3bdd8-803e-4985-8823-932586fa7ca1
                © The Author(s) 2019

                Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

                History
                : 11 September 2018
                : 19 February 2019
                Funding
                Funded by: FundRef https://doi.org/10.13039/501100001691, MEXT | Japan Society for the Promotion of Science (JSPS);
                Award ID: JP16K12871
                Award ID: JP18K19912
                Award Recipient :
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