Measurement of Spatiotemporal Intracellular Deformation of Cells Adhered to Collagen Matrix During Freezing of Biomaterials

Abstract

Preservation of structural integrity inside cells and at cellextracellular matrix (ECM) interfaces is a key challenge during freezing of biomaterials. Since the postthaw functionality of cells depends on the extent of change in the cytoskeletal structure caused by complex cellECM adhesion, spatiotemporal deformation inside the cell was measured using a newly developed microbeadmediated particle tracking deformetry (PTD) technique using fibroblastseeded dermal equivalents as a model tissue. Fibronectincoated 500 nm diameter microbeads were internalized in cells, and the microbeadlabeled cells were used to prepare engineered tissue with type I collagen matrices. After a 24 h incubation the engineered tissues were directionally frozen, and the cells were imaged during the process. The microbeads were tracked, and spatiotemporal deformation inside the cells was computed from the tracking data using the PTD method. Effects of particle size on the deformation measurement method were tested, and it was found that microbeads represent cell deformation to acceptable accuracy. The results showed complex spatiotemporal deformation patterns in the cells. Large deformation in the cells and detachments of cells from the ECM were observed. At the cellular scale, variable directionality of the deformation was found in contrast to the onedimensional deformation pattern observed at the tissue scale, as found from earlier studies. In summary, this method can quantify the spatiotemporal deformation in cells and can be correlated to the freezinginduced change in the structure of cytosplasm and of the cellECM interface. As a broader application, this method may be used to compute deformation of cells in the ECM environment for physiological processes, namely cell migration, stem cell differentiation, vasculogenesis, and cancer metastasis, which have relevance to quantify mechanotransduction.