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Research Papers

Role of Cytoskeletal Components in Stress-Relaxation Behavior of Adherent Vascular Smooth Muscle Cells

[+] Author and Article Information
Jason D. Hemmer1

Department of Bioengineering, 401 Rhodes Engineering Research Center, Clemson University, Clemson, SC 29634jhemmer@clemson.edu

Jiro Nagatomi, Scott T. Wood, Alexey A. Vertegel, Delphine Dean, Martine LaBerge

Department of Bioengineering, 401 Rhodes Engineering Research Center, Clemson University, Clemson, SC 29634

1

Corresponding author. Present address: 401 Rhodes Engineering Research Center, Clemson University, Clemson, SC 29634.

J Biomech Eng 131(4), 041001 (Jan 20, 2009) (9 pages) doi:10.1115/1.3049860 History: Received May 01, 2008; Revised October 14, 2008; Published January 20, 2009

A number of recent studies have demonstrated the effectiveness of atomic force microscopy (AFM) for characterization of cellular stress-relaxation behavior. However, this technique’s recent development creates considerable need for exploration of appropriate mechanical models for analysis of the resultant data and of the roles of various cytoskeletal components responsible for governing stress-relaxation behavior. The viscoelastic properties of vascular smooth muscle cells (VSMCs) are of particular interest due to their role in the development of vascular diseases, including atherosclerosis and restenosis. Various cytoskeletal agents, including cytochalasin D, jasplakinolide, paclitaxel, and nocodazole, were used to alter the cytoskeletal architecture of the VSMCs. Stress-relaxation experiments were performed on the VSMCs using AFM. The quasilinear viscoelastic (QLV) reduced-relaxation function, as well as a simple power-law model, and the standard linear solid (SLS) model, were fitted to the resultant stress-relaxation data. Actin depolymerization via cytochalasin D resulted in significant increases in both rate of relaxation and percentage of relaxation; actin stabilization via jasplakinolide did not affect stress-relaxation behavior. Microtubule depolymerization via nocodazole resulted in nonsignificant increases in rate and percentage of relaxation, while microtubule stabilization via paclitaxel caused significant decreases in both rate and percentage of relaxation. Both the QLV reduced-relaxation function and the power-law model provided excellent fits to the data (R2=0.98), while the SLS model was less adequate (R2=0.91). Data from the current study indicate the important role of not only actin, but also microtubules, in governing VSMC viscoelastic behavior. Excellent fits to the data show potential for future use of both the QLV reduced-relaxation function and power-law models in conjunction with AFM stress-relaxation experiments.

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Copyright © 2009 by American Society of Mechanical Engineers
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Figures

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Figure 2

Sample indentation curve illustrating approximate probe-cell separation distance. Force values were not calculated for indentation curves used in the determination of separation distance.

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Figure 1

Sample stress-relaxation and Z sensor-strip chart data. The top chart (channel 1) displays cantilever deflection force versus time. Deflection force has been calibrated and shifted to a baseline of 0 nN. The bottom chart (channel 2) displays Z-piezo movement versus time.

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Figure 3

Averaged normalized VSMC relaxation curves plotted versus logarithmic time from (a) cytochalasin D, (b) jasplakinolide, (c) paclitaxel, and (d) nocodazole. The arrows indicate the direction that curves of treated cells are shifted relative to controls. Due to the usage of logarithmic spacing and the 100 Hz initial sampling rate, the initial time point of each curve is 0.01 s.

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Figure 4

Representative example of a normalized VSMC relaxation curve plotted versus logarithmic time with curve fits of (a) QLV reduced-relaxation, (b) power-law, and (c) SLS models. Due to the usage of logarithmic spacing and the 100 Hz initial sampling rate, the initial time point of each curve is 0.01 s.

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Figure 5

Immunofluorescence images of untreated VSMC (a) actin and (b) microtubules; cytochalasin D-treated VSMC (c) actin and (d) microtubules; nocodazole-treated VSMC (e) actin and (f) microtubules; paclitaxel-treated VSMC (g) actin and (h) microtubules; jasplakinolide-treated VSMC (i) actin and (j) microtubules. Scalebars represent 50 μm.

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