Abstract
Interaction of integrins with extracellular matrices is essential for cell adhesion to substrata. Ventral surfaces of fibroblasts adhering to flat substrata are not flat but have uneven 3D topology. However, spatial relationship between the topology of the ventral cell surface and arrangement of extracellular matrix fibrils remains unclear. Here, we report a novel and simple method based on total internal reflection fluorescence microscopy to quantify the distance between the ventral plasma membrane and the glass substratum. We observe that the distance varies from < 25 nm at focal adhesions to 40–50 nm at close contacts and > 80 nm in other regions. Furthermore, by applying this novel method, we show that fibronectin fibrils are also separated from the substratum in regions where the ventral cell surface-substratum distance is > 80 nm. Our results reveal that fibronectin fibrils are not simply adsorbed to the glass substratum but follow the ventral cell surface topology.
ACKNOWLEDGEMENTS
We thank Dr. Sri Ram Krishna Vedula (National University of Singapore) for proofreading of the manuscript and Dr. Hiroaki Machiyama (National University of Singapore) for technical support. We also thank two reviewers’ helpful comments that significantly improved the manuscript. This work was supported in part by Grants-in-aid for General Scientific Research from the Ministry of Education Science Sports and Culture (to HM).
Declaration of interest: The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the article.