Each year more than 45,000 Americans suffer burns serious enough to require a hospital stay, according to the American Burn Association. While the traditional therapy of using skin grafts to cover burn sites has improved, a number of problems including scarring, infection and poor adhesion remain.
“Skin grafts involve taking skin (both the upper epidermal and the underlying dermis) from an unburned site on the patient’s body or from a cadaver and grafting it on to the burn wound,” said Craig D. Woodworth, a cell biologist and associate professor at Clarkson University. “Skin grafts often require multiple surgeries. Cadaver skin is scarce and can introduce disease. In the case of extensive burns, large amounts of skin can be created by isolating individual epidermal cells and then expanding their numbers in culture, but the skin simply does not look or function like normal skin. There are no hair follicles, no pores for sweating, and the pigment is often a poor match.”
Woodworth is collaborating with Anja Mueller, a polymer chemist and assistant professor of chemistry at Clarkson, on research to develop an artificial skin that would heal and function like normal skin and could be used successfully for large burns or surgical reconstruction.
“Our goal is to bioengineer an artificial skin scaffold that promotes tissue regeneration and even directs cell growth for hair follicles and sweat glands so that the new skin would look and feel like normal skin,” said Woodworth.
Though still in its early stages, their initial research looks promising. While Woodworth focuses on isolating a combination of cytokines that will generate skin growth and promote wound healing, Mueller is creating and testing biodegradable polymers to find one that will support cell growth and regeneration.
Cytokines are naturally occurring proteins that regulate or modify the growth of specific cells. The development of skin is complex and there are hundreds of different cytokines that are made by skin cells. However, only a subset is critical for skin growth and differentiation.
“An important question, then, is which cytokines are most effective in helping the graft heal and restore normal function,” explained Woodworth. “We are focusing on several, including EGF (epidermal growth factor) that stimulates cells to grow quickly and fill in the wound and VEGF (vascular endothelial growth factor) that stimulates blood vessel formation and nourishes the grafted skin.”
Recently, Woodworth has been working in the laboratory analysing the effects of Transforming Growth Factor-B1 (TGF-B1) on gene expression in skin cells. This cytokine regulates production of certain cell proteins, namely collagen and connective tissue proteins, associated with wound healing.
“Our results show that TGF-B1 stimulates the formation of collagen, the natural scaffold for skin, by the fibroblasts, the underlying supporting cells,” explained Woodworth. “However, long-term expression of TGF-B1 also contributes to scarring when it is produced after the initial healing has occurred. Therefore, the trick may be to turn TGF-B1 on initially, but then turn if off when the skin begins to restore normal structure and function.”
Meanwhile, in her laboratory Mueller is testing biodegradable polymers in order to find the “right” polymer that will maximise the function of a bioengineered skin scaffold.
“There are two major issues. The first one is to develop a polymer that cells really like to attach to and grow on,” said Mueller. “For example, surface structure, charge and degree of cross linking of the polymer may all affect cell behaviour. The second challenge is to develop a polymer that can undergo slow degradation in the body, and allow the transplanted cells to start making their own collagen and basement membrane that would rebuild the natural skin scaffold. The polymer scaffold might be considered then a temporary structure to provide the initial support of new skin.”
Just as important, according to Mueller, the polymer should be designed to slowly release the cytokines that stimulate the reorganisation of natural skin. This is where a mild enzymatic synthesis under development by Mueller has an advantage.
Her approach, which involves incorporating the cytokines into the polymer scaffold before it is assembled, is a novel one. According to Mueller, this method allows for a steadier, more controlled release of the cytokines than the traditional method of adding the cytokines later in the process after the scaffold is assembled. Her method of synthesis also does not require a toxic catalyst or organic solvents so there is no danger of residual toxic compounds in the skin scaffold.
The collaborators will explore the complex problem of skin differentiation (such as how to organise glands, hair follicles, proper pigment cells) and the appropriate mix of cytokines by making skin in a cell culture dish where it can be more easily studied.
“Most scientists grow isolated skin cells as a flat layer on the bottom of a culture dish, submerged in cell culture medium,” explained Woodworth. “However, if you add the appropriate cells and scaffold, then grow the cells at the interface between the culture media and the air (on top of the culture medium), the system spontaneously forms a multilayered skin, in a way that is similar to what occurs naturally.”
Woodworth and Mueller will use this organotypic culture system to test for the most effective mix of cytokines and polymers. It has the added advantage that the scientists would not need to use experimental animals to develop the proper system.
“Ultimately, we are interested in understanding and finding the right mixture of biological and synthetic materials that will yield the best results for wound healing and tissue regeneration and will significantly increase the quality of life for severe burn victims and medical patients.”