Biomedical engineers at Washington University in St. Louis have developed new biomaterials to recruit endothelial cells to the inner surfaces of vascular grafts. Endothelial cells normally line blood vessels and actively protect against blood clotting. Blood clotting on artificial materials is currently so severe that the use of vascular grafts is limited to large diameter vessels.
A team led by Donald Elbert, Ph. D., Washington University assistant professor of biomedical engineering, synthesized the new materials. The materials are about 50 per cent synthetic polymers and 50 per cent protein. The polymer portion of the materials is a derivative of polyethylene glycol that was initially synthesized by Washington University graduate student Evan A. Scott. When a solution of the polymer is mixed with protein at the proper ratio, a chemical reaction leads to the formation of a water-swollen hydrogel.
The materials perform a variety of functions – limiting protein activation, providing cell adhesion cues to the endothelial cells and delivering molecules that enhance endothelial cell migration and survival. The polymer portion limits the activation of blood clotting proteins normally associated with artificial materials, while the protein portion traps a signaling molecule that promotes endothelial cell migration and survival. Endothelial cells grow on the surface of the materials due to the presence of chemically synthesized molecules that specifically bind to adhesion receptors on the cell surface.
In a study published recently in the journal Biomacromolecules, graduate student Bradley K. Wacker demonstrated that the migration speed of endothelial cells on the materials doubles when the signaling lipid sphingosine 1-phosphate (S1P) is delivered from the protein part of the material. S1P is a small molecule that is used for intercellular communication in the body. It interacts with receptors on endothelial cells to promote migration and survival, while limiting the migration of cells in the middle portion of the artery that sometimes cause narrowing of blood vessels.
“The challenges were unique, since the need for a lipid delivery system hadn’t been previously considered,” said Elbert.
The work is supported by a $1,493,744 grant from the National Institutes of Health.
Lipid delivery system
Sphingosine 1-phosphate is critical in the development of the vascular system, but it is also active in other parts of the body. For example, it is critical for the maturation of lymphocytes. Thus, Elbert and colleagues are working to localize the effects of the potent signaling lipid. The lipid is released over time from the materials but the endothelial cells sitting on the materials are the first cells to see the lipid. Afterwards, the lipid is quickly diluted and its effects are minimized. For example, Wacker can place two small pieces of the materials in the same cell culture dish, where one piece is loaded with the signaling lipid and the other is not. Only the cells on the lipid-releasing material display an increase in cell migration rates.
The materials also may be useful to promote angiogenesis, the process by which new blood vessels are formed from the existing vasculature. Angiogenesis is a vital part of most tissue-engineering strategies to allow delivery of oxygen and nutrients to developing tissues. If the new tissue is not vascularized, ischemia, or a lack of blood supply, will result in cell death and a loss of tissue mass. The implantation of biomaterials to promote angiogenesis is also an emerging therapeutic strategy for the treatment of ischemia that results from heart disease or peripheral vascular disease.
Angiogenesis in adults occurs mainly at the sites of injury, mediated by the release of a variety of factors in response to low levels of oxygen. Part of the process of angiogenesis involves the local breakdown of the existing vasculature. This leads to localized blood clotting and the release of the lipid factor S1P. S1P stimulates endothelial cell migration, proliferation, and entubulation, leading to new blood vessel formation. Wacker tested the S1P-releasing hydrogels using a test called a chorioallantoic membrane assay, which allowed him to directly observe the extent to which new blood vessels formed in response to controlled delivery of the lipid.
The delivery of S1P from these materials may prove to be particularly useful in conjunction with other factors that are currently being studied for therapeutic angiogenesis. Graduate student Shannon Alford has found that the effect of S1P delivery might be amplified by other stimuli such as shear stress or vascular endothelial growth factor. One particularly promising avenue is to synthesize the active S1P following implantation, by immobilizing within the material the enzyme that normally produces S1P within the body. Biomedical engineering graduate student Megan Kaneda first cloned and expressed this enzyme in a manner that allows its incorporation within the material.
“This is a really unique opportunity in the field of drug delivery because we aren’t limited by the amount of drug that can be loaded in the material,” Elbert said. “We are able to take advantage of the fact that the precursor to S1P is already present in the bloodstream.”