Title

The Culture of Dermal Replacements

Document Type

Thesis

Publication Date

1997

Advisor

Elizabeth Wurdak

Abstract

Collagen-based wound dressings have been used to prevent excessive fluid loss and infection in individuals with severe skin damage, as well as to promote the re-growth and healing of the patient's own skin. In this study, problems encountered with cryopreservation of collagen-based wound dressings were researched.

A fibroblast cell-line was expanded in culture and seeded onto collagen sponges. The sponges were then cultured for varying lengths of time before freezing them at -80 degrees C. The sponges were analyzed to assess the total number of cells as a function of time in culture.

Toxicity tests were performed using varying concentrations of two cryoprotectants, Simethyl Sulfoxide (DMSO) and Glycerol. Varying concentrations were tested in an effort to find the concentration of each that gave the highest cell survival and minimal toxic effects.

Over a four hour period, concentrations of up to 10% DMSO did not have an effect on cell viability. The results obtained at 12% DMSO showed that the viability of the cells began to decrease at three hours, and 15% DMSO had a gradual, increasingly toxic effect over time. At glycerol concentrations of 5, 10, and 15%, it was observed that the total cell number was reduced.

Efforts were made to culture sponges to obtain a concentration of 1 x 10 (9) fibroblast cells per sponge in order to provide a detectable signal for analysis by magnetic Resonance Imaging. Using MRI, the mechanism of cell death due to cryopreservants can be observed and a more efficient protocol for storing sponges created. However, the highest number of cells cultured per sponge was less than 1.4 x 10 (8). Additional research is needed to improve either the fibroblast concentration on collagen sponges or the resolution obtainable by MRI analysis.

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