(A-29) In-situ observation of dynamic nuclear deformation induced by cell-substrate stretching

Introduction:: Nuclear deformation caused by external mechanical stimuli has been suggested to impact cellular activities, such as cell migration, gene expression, and proliferation; however, the underlying mechanisms remain largely unknown. Since cells reside in dynamic mechanical environments, investigation of viscoelastic behaviors of nuclei under mechanical conditions is believed to be a key to understanding the mechanisms of cellular responses to mechanical stimuli. In this study, we developed an in-situ observation system for the dynamic deformation behavior of the nucleus under mechanical substrate stretching conditions.

Materials and Methods:: Human dermal fibroblasts cultured in a custom-made PDMS chamber were subjected to 5% stretch at a low (0.005/s) or high (0.025/s) strain rate by using a newly developed stretching system equipped on the stage of an inverted microscope. To obtain sequential "in-focus" fluorescent images of the nucleus during cell stretching conditions, the height of the stretching chamber was continuously controlled with a polynomial interpolation method during image acquisition. Intranuclear strains were then calculated from the fluorescence images obtained. Fluorescence images of the nucleus before and during stretching were applied to a 2D digital image correlation program to calculate local displacements and Green-Lagrangian strain within the nucleus.

Results, Conclusions, and Discussions:: We successfully obtained in-focused nuclear fluorescent images of fibroblast nuclei at several strain points during substrate stretching and time course changes of intranuclear strain. Nuclear strain caused by cell-substrate stretching increased in parallel and decreased in perpendicular to the direction of stretching regardless of strain rate. However, compared to the low strain rate condition, the whole nuclear strain in the direction of stretching tends to decrease during the application of stretching at the high strain rate. In addition, the intranuclear strain showed heterogeneous magnitude distribution under the high strain rate condition, while a uniform strain distribution was observed at the low strain rate. These strain rate-dependent deformation behaviors are thought to reflect viscoelastic mechanical properties of not only the whole nucleus but also local intranuclear structures such as chromatin and intranuclear lamin structures. The system we developed will help reveal the role of viscoelastic behaviors of intranuclear structures in the cellular response to substrate stretching.

Acknowledgements (Optional): : This study was supported in part byGrants-in-Aid for Scientific Research from the MEXT of Japan (Nos. 18K19934, and 22K19898), a Tokyo Metropolitan Government Advanced Research Grant (R2-2), and the TMU Research Project for Emergent Future Society.


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