Design Innovation Papers

Three-Dimensional Culture of Cells and Matrix Biomolecules for Engineered Tissue Development and Biokinetics Model Validation

[+] Author and Article Information
Shelley S. Mason, Randy D. Zelick

Regenerative Bioengineering Laboratory, Department of Mechanical and Materials Engineering, and Department of Biology, Portland State University, Portland, OR 97201

Sean S. Kohles1

Regenerative Bioengineering Laboratory, Department of Mechanical and Materials Engineering, and Department of Biology, Portland State University, Portland, OR 97201kohles@cecs.pdx.edu

Shelley R. Winn

Department of Molecular and Medical Genetics, Oregon Health & Science University, Portland, OR 97239

Asit K. Saha

Center for Allaying Health Disparities Through Research and Education (CADRE), Department of Mathematics and Computer Science, Central State University, Wilberforce, OH 45384


Corresponding author.

J. Nanotechnol. Eng. Med 2(2), 025001 (May 19, 2011) (7 pages) doi:10.1115/1.4003878 History: Received March 13, 2011; Revised March 23, 2011; Published May 19, 2011; Online May 19, 2011

There has been considerable progress in cellular and molecular engineering due to recent advances in multiscale technology. Such technologies allow controlled manipulation of physiochemical interactions among cells in tissue culture. In particular, a novel chemomechanical bioreactor has recently been designed for the study of bone and cartilage tissue development, with particular focus on extracellular matrix formation. The bioreactor is equally significant as a tool for validation of mathematical models that explore biokinetic regulatory thresholds (Saha, A. K., and Kohles, S. S., 2010, “A Distinct Catabolic to Anabolic Threshold Due to Single-Cell Nanomechanical Stimulation in a Cartilage Biokinetics Model,” J. Nanotechnol. Eng. Med., 1(3), p. 031005; 2010, “Periodic Nanomechanical Stimulation in a Biokinetics Model Identifying Anabolic and Catabolic Pathways Associated With Cartilage Matrix Homeostasis,” J. Nanotechnol. Eng. Med., 1(4), p. 041001). In the current study, three-dimensional culture protocols are described for maintaining the cellular and biomolecular constituents within defined parameters. Preliminary validation of the bioreactor’s form and function, expected bioassays of the resulting matrix components, and application to biokinetic models are described. This approach provides a framework for future detailed explorations combining multiscale experimental and mathematical analyses, at nanoscale sensitivity, to describe cell and biomolecule dynamics in different environmental regimes.

Copyright © 2011 by American Society of Mechanical Engineers
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Figure 1

Three-dimensional reconstruction via microcomputed tomography of a through-thickness section of human articular cartilage (after (40)). The total transverse slice dimension is 1500 μm representing the articular surface (top), three histologic zonal layers, and the bony osteochondral transition (bottom).

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Figure 2

A solid model of the assembled chemomechanical bioreactor for three-dimensional culture (after (22)). The image includes the upper electromechanical actuator applying high-resolution load-deformation control to the “hat,” which distributes the mechanical stimulus to the five load arms and, as such, to the five culture wells below. The wells can be rotated into the view of single ultrasonic or digital imaging sensors.

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Figure 3

The now-fabricated, novel chemomechanical bioreactor with (a) five culture wells and (b) a load-train applying load-displacement for measuring exact loads. The high-resolution, copper-colored load-cells measure the dynamic loads transferred via the load-rods through the square load-platens into the culture wells and onto the eventual cell-biomaterial constructs.

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Figure 4

Open-view (model) of a bioreactor test chamber for engineered tissue chemomechanical stimulation indicating the (a) cell-biomaterial construct (125 mm3 pink cube) and (b) three current well designs for static and dynamic three-dimensional culture. The middle culture well, which provides perfused flow and dynamic culture, will be described in a future article (21).

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Figure 5

Expected cell-matrix preparations (after (25)). During matrix synthesis, cells will be distributed throughout the (a) collagen (to be stained with Masson’s Trichome) and (b) aggrecan (to be stained with safranin-O/fast) constituents as are typical for this cell-scaffold arrangement. The solid dark circles each represent a single cell at ∼20 μm in diameter.

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Figure 6

Representative biokinetic modeling results (applying Eq. 6) from expected biomolecular accumulation data as gathered from the chemomechanical bioreactor (as boundary conditions and validation). Here the model is initially allowed to run as the ECM concentration (%, GAG and collagen) peaks in the response to local resources and then drops to a steady state level. A chemomechanical stimulus is then applied at 5×105 relative time units, driving the accumulation upward in an anabolic response.




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