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Technical Briefs

Controlled Vacuum Seeding as a Means of Generating Uniform Cellular Distribution in Electrospun Polycaprolactone (PCL) Scaffolds

[+] Author and Article Information
Ming Chen

Biomedical Engineering and Biotechnology Program, University of Massachusetts Dartmouth, North Dartmouth, MA 02747

Heather Michaud

Department of Materials and Textiles, University of Massachusetts Dartmouth, North Dartmouth, MA 02747

Sankha Bhowmick1

Biomedical Engineering and Biotechnology Program, Department of Mechanical Engineering, University of Massachusetts Dartmouth, North Dartmouth, MA 02747sbhowmick@umassd.edu

1

Corresponding author.

J Biomech Eng 131(7), 074521 (Jul 27, 2009) (8 pages) doi:10.1115/1.3173283 History: Received October 15, 2008; Revised May 29, 2009; Published July 27, 2009

A major challenge encountered in using electrospun scaffolds for tissue engineering is the non-uniform cellular distribution in the scaffold with increasing depth under normal passive seeding conditions. Because of the small surface pores, typically few microns in diameter, cells tend to congregate and proliferate on the surface much faster compared to penetrating the scaffold interior. In order to overcome this problem, we used a vacuum seeding technique on polycaprolactone electrospun scaffolds while using NIH 3T3 fibroblasts as the model cell system. This serves as a precursor to the bilayer skin model where the fibroblasts would be residing at an intermediate layer and the keratinocytes would be on the top. Vacuum seeding was used in this study to enhance fibroblasts seeding and proliferation at different depths. Our results show that the kinetics of cell attachment and proliferation were a function of varying vacuum pressure as well as fiber diameter. Cell attachment reached a maxima somewhere between 2–8 in. Hg vacuum pressure and fell for lower vacuum pressures presumably because of cell loss through the filtration process. Cell proliferation and collagen secretion over five days indicated that vacuum pressure did not affect cellular function adversely. We also compared the combined impact of scaffold architecture (400 nm versus 1100 nm average diameter fiber scaffolds) and vacuum pressure. At a given pressure, more cells were retained in the 400 nm scaffolds compared to 1100 nm scaffolds. In addition, the cell intensity profile shows cell intensity peak shift from the top to the inner layers of the scaffold by lowering the vacuum pressure from 0 in. Hg to 20 in. Hg. For a given vacuum pressure the cells were seeded deeper within the 1100 nm scaffold. The results indicate that cells can be seeded in electrospun scaffolds at various depths in a controlled manner using a simple vacuum seeding technique. The depth of seeding is a function of pressure and scaffold fiber diameter.

FIGURES IN THIS ARTICLE
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Copyright © 2009 by American Society of Mechanical Engineers
Topics: Vacuum , Pressure
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Figures

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Figure 1

Schematic of the vacuum seeding setup used for the experiment in this study

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Figure 2

(a) Surface pore diameter distribution on the 400 nm PCL electrospun scaffolds, and (b) Surface pore diameter distribution on the 1100 nm PCL electrospun scaffolds

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Figure 3

Cell attachment as a function of vacuum pressure on the 400 nm and 1100 nm scaffolds at 4 h and 8 h

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Figure 4

Cell proliferation as a function of vacuum pressure on the 400 nm and 1100 nm scaffolds

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Figure 5

Collagen secretion in the 400 nm and 1100 nm scaffolds under different vacuum pressures

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Figure 6

Cell spatial distribution under different vacuum seeding pressures: (a) typical confocal images in different layers; (b) cell intensity in the 400 nm scaffolds at day 3; (c) cell intensity in the 1100 nm scaffolds at day 3

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