Research Papers

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

[+] Author and Article Information
Soham Ghosh

School of Mechanical Engineering,
Purdue University,
West Lafayette, IN 47906

J. Craig Dutton

Department of Aerospace Engineering,
University of Illinois at Urbana-Champaign,
Urbana, IL 61801

Bumsoo Han

School of Mechanical Engineering,
Weldon School of Biomedical Engineering,
Purdue University,
West Lafayette, IN 47906
e-mail: bumsoo@purdue.edu

1Corresponding author.

Contributed by the Bioengineering Division of ASME for publication in the Journal of Biomechanical Engineering. Manuscript received September 15, 2013; final manuscript received December 2, 2013; accepted manuscript posted December 9, 2013; published online February 5, 2014. Editor: Victor H. Barocas.

J Biomech Eng 136(2), 021025 (Feb 05, 2014) (8 pages) Paper No: BIO-13-1430; doi: 10.1115/1.4026180 History: Received September 15, 2013; Revised December 02, 2013; Accepted December 09, 2013

Preservation of structural integrity inside cells and at cell-extracellular matrix (ECM) interfaces is a key challenge during freezing of biomaterials. Since the post-thaw functionality of cells depends on the extent of change in the cytoskeletal structure caused by complex cell-ECM adhesion, spatiotemporal deformation inside the cell was measured using a newly developed microbead-mediated particle tracking deformetry (PTD) technique using fibroblast-seeded dermal equivalents as a model tissue. Fibronectin-coated 500 nm diameter microbeads were internalized in cells, and the microbead-labeled 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 one-dimensional 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 freezing-induced change in the structure of cytosplasm and of the cell-ECM 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.

Copyright © 2014 by ASME
Your Session has timed out. Please sign back in to continue.



Grahic Jump Location
Fig. 1

(a) Triangular element deforms from an initial (t = 0) to a later configuration (at time t) with accompanied translation and rotation. The initial and final configurations are used to compute the deformation gradient tensor. (b) Regional deformation analysis scheme to delineate deformation from translation and rotation. Deformation gradient tensor F, left Cauchy-Green tensor B and finally the stretch ratios λ1 and λ2 are computed sequentially during the method. The stretch ratios are used to compute the first invariant I1 and second invariant I2.

Grahic Jump Location
Fig. 2

Imposed temperature history characterization at location of interest. Temperature history for the present study was matched (n = 3) with the average temperature history at x = 2 mm, performed with a traditional directional freezing stage [23].

Grahic Jump Location
Fig. 3

(a) Fluorescent microbead-labeled cell attached on polymerized collagen gel. The dotted line shows the location of the freeze front at a given time as it propagates gradually from left towards right. (b) Triangular meshes have been generated using the Delaunay algorithm using the tracks of the microbeads. Some triangles are rejected based on the criteria stated in the main text. The microbeads are tracked during postprocessing after the cell deformation experiment.

Grahic Jump Location
Fig. 4

Movement of nanoparticles inside the cell. For microbeads the relative position of the particles remains relatively constant (two representative particles are marked by arrows) over a 2 h time interval as shown in (a). For QDs (two representative particles are marked by arrows) the relative positions between particles change over time as shown in (b). Loci of the two representative particles over 2 h are presented in (c) for microbeads and (d) for QDs. The initial positions (at t = 0) of the nanoparticles are marked by the arrows. For microbeads, the loci are pathlines of two particles although they look like points implying very small movement of the particles. For the same representative pair of nanoparticles, the normalized inter-particle distance r(t)/r(t = 0) with time is presented in (e). The value remains close to 1 for microbeads but varies substantially for QDs. The same result is observed for inter-particle angular orientation φ(t)/φ(t = 0) between particles (f). This behavior was observed for several pairs (n = 10) of particles of both sizes.

Grahic Jump Location
Fig. 5

Deformation in two representative cells (a) and (b) cultured on collagen gel. Arrow in (i) shows the direction of freeze front propagation. From the fluorescence images (i), (ii), and (iii) the cell deformation can be visualized. The surface plot (iv) shows the spatial distribution of I1 at a given intermediate time. Surface plot for I2 is presented in (v) at the same intermediate time. For the same time point, the directionality of the stretch ratios is presented in (vi) for some triangular elements. The pair of arrows represent mutually perpendicular principal stretch ratios (magnitude and directions found as eigenvalues and eigenvectors of left Cauchy-Green tensor B). The scale bar represents the unit stretch ratio with reference to the size of the arrows. Spatially they are variably oriented in the cell and different in magnitude from one part of the cell to the other.

Grahic Jump Location
Fig. 6

Spatiotemporal intracellular deformation for a representative cell cultured on collagen gel. The top panel in (a) shows the deforming cell with internalized fluorescent microbeads. The arrow at bottom of the 3 s image shows the direction of freeze front propagation. Bottom panel in (a) shows the surface plot of I1 for the same time points. The individual temporal evolution of I1 for the three triangular elements indicated by b, c, d in (a) are presented in (b), (c), and (d).




Some tools below are only available to our subscribers or users with an online account.

Related Content

Customize your page view by dragging and repositioning the boxes below.

Related Journal Articles
Related eBook Content
Topic Collections

Sorry! You do not have access to this content. For assistance or to subscribe, please contact us:

  • TELEPHONE: 1-800-843-2763 (Toll-free in the USA)
  • EMAIL: asmedigitalcollection@asme.org
Sign In