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A Compressed Collagen Construct for Studying Endothelial–Smooth Muscle Cell Interaction Under High Shear Stress

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Abstract

The coculture of vascular endothelial cells (ECs) on collagen gels containing smooth muscle cells (SMCs) has been carried out to investigate cellular interactions associated with blood vessel pathophysiology under wall shear stress (WSS) conditions. However, due to a lack of gel stiffness, the previous collagen gel coculture constructs are difficult to use for pathologic higher WSS conditions. Here, we newly constructed a coculture model with centrifugally compressed cell–collagen combined construct (C6), which withstands higher WSS conditions. The elastic modulus of C6 was approximately 6 times higher than that of the uncompressed collagen construct. The level of α-smooth muscle actin, a contractile SMC phenotype marker observed in healthy arteries, was elevated in C6 compared with that of the uncompressed construct, and further increased by exposure to a physiological level WSS of 2 Pa, but not by a pathological level of 20 Pa. WSS conditions of 2 and 20 Pa also induced different expression ratios of matrix metalloproteinases and their inhibitors in the C6 coculture model but did not in monocultured ECs and SMCs. The C6 coculture model will be a powerful tool to investigate interactions between ECs and SMCs under pathologically high WSS conditions.

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Acknowledgments

The present study was supported in part by Grants-in-Aid for Scientific Research from the MEXT of Japan (Grant Nos. 17K10763, 18K08770, 18K19934, 20K09154, and 21K08828) and Tokyo Metropolitan Government Advanced Research Grant (Grant No. R2-2).

Funding

Funding was provided by Ministry of Education, Culture, Sports, Science and Technology and Tokyo Metropolitan Government (Grant No. R2-2).

Conflict of interest

The authors declare that they have no conflicts of interest.

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Authors and Affiliations

  1. Department of Cardiovascular Surgery, Jichi Medical University, Yakushiji 3311-1, Shimotsuke, Tochigi, 329-0498, Japan

    Yuya Hiroshima & Koji Kawahito

  2. Department of Mechanical Systems Engineering, Tokyo Metropolitan University, Minami-Osawa 1-1, Hachioji, Tokyo, 192-0397, Japan

    Yuki Oyama, Kaoru Sawasaki, Hiromichi Fujie & Naoya Sakamoto

  3. Department of Electrical and Mechanical Engineering, Nagoya Institute of Technology, Gokiso, Showa, Nagoya, 466-8555, Japan

    Masanori Nakamura

  4. Department of Cardiovascular Surgery, Saitama Medical Center, Jichi Medical University, Amanuma 1-847, Omiya, Saitama, 330-834, Japan

    Naoyuki Kimura

  5. Research Center for Medicine-Engineering Collaboration, Tokyo Metropolitan University, Minami-Osawa 1-1, Hachioji, Tokyo, 192-0397, Japan

    Hiromichi Fujie & Naoya Sakamoto

Authors
  1. Yuya Hiroshima
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  2. Yuki Oyama
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Correspondence to Naoya Sakamoto.

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Associate Editor Jane Grande-Allen oversaw the review of this article.

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Supplementary Information

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Supplementary file1 (TIFF 10263 KB)

Supplementary Figure S1 Relative expressions of MMP-2, MMP-9, TIMP-1, and TIMP-2 of SMCs in compressed collagen constructs. Results are normalized by GAPDH and expressed as relative to the expression of SMCs in the uncompressed collagen constructs. *p < 0.05 (Mann-Whitney U test).

Supplementary file2 (TIFF 10263 KB)

Supplementary Figure S2 Representative fluorescence images of calponin-1 of SMCs in the uncompressed (left) and the compressed (C6)(right) collagen constructs. Bars = 100 µm. With a primary antibody for calponin-1 (1:200, Cell Signaling Technology), fluorescent staining was performed according to the method stated in the manuscript. By counting the number of cells in the images, we found the ratio of calponin-1 expressing cells in the compressed (C6) construct was 0.29 ± 0.02, while that in the uncompressed gel was 0.23 ± 0.02 (mean ± SEM, n = 3).

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Hiroshima, Y., Oyama, Y., Sawasaki, K. et al. A Compressed Collagen Construct for Studying Endothelial–Smooth Muscle Cell Interaction Under High Shear Stress. Ann Biomed Eng 50, 951–963 (2022). https://doi.org/10.1007/s10439-022-02972-7

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  • DOI: https://doi.org/10.1007/s10439-022-02972-7

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